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Full text of "Manual of veterinary microbiology;"

/.nT;it CI it iu> 11,30 

MANUAL 



VETERINARY MICROBIOLOGY. 



(^PROFESSOBS)^OSSELM AN AND ^LIENAUX, 

National Veterinary College, Cureghem, Belgium. 



TRANSLATED AND EDITED BY 

R. R DINWIDDIE, 

Professor of Veterinary Science, College of Agriculture, Arkansas State Uni- 
versity; Animal Pathologist, Arkansas Agricultural Experiment Station. 




CINCINNATI: 

THE ROBERT CLARKE COMPANY. 
1894. 



COPYRIGHT, 1894, 
BY R. R. DINWIDDIE 



PREFACE. 



In undertaking the translation of MM. Mosselman 
and Lienaux's "Manuel de Microbioloyie Veterinaire," . 
my object has been to supply English speaking veteri- 
nary students and practitioners with a work on Bac- 
teriology which seemed specially adapted to their 
needs. The book is small, but it conveys more in- 
formation on the etiology of the infectious diseases 
of animals and the biology of the germs associated 
with them than any other single work in our lan- 
guage. Unlike other works on Bacteriology accessi- 
ble to readers of English only, the Microbiology of 
animal diseases is treated of as the essential part of 
the work, that of diseases of mankind only incident- 
ally referred to. The completeness and accuracy as 
to details with which it discusses the modes of prop- 
agation of some of our most important diseases and 
the general conditions under which these diseases 
occur, ought to recommend the book to practical 
veterinarians, who are presumably more interested in 
the ascertained facts in regard to any disease than in 
the individuality of the germ which occasions it. 

The book is not intended for a laboratory manual, 
consequently, the technique of staining and cultiva- 

(iii) 



iv Preface. 

tion of germs is not exhaustively discussed, and the 
usual illustrations of bacteriological apparatus have 
been omitted. 

As to the translation itself a few words are neces- 
sary : Weights and measures, given in the metrical 
system in the original, have not been changed. Di- 
mensions which occur in this work are chiefly those 
of microscopic objects which are now rarely ex- 
pressed by American microscopists in fractions of an 
inch. The thermometric readings are in all cases 
given in the centigrade scale. For those who are 
unfamiliar with the decimal system the Appendix 
will supply the requisite information. 

The few foot-notes which I have introduced are in 
some cases intended to be supplementary to the text, 
referring to discoveries which have been made since 
the publication of the original in 1891. In other 
cases they are explanatory of words or statements 
which might otherwise be misunderstood. 

To the illustrations which occur in the French 
text I have added a few others borrowed from differ- 
ent sources, which are acknowledged in the descrip- 
tions accompanying the figures ; four are from draw- 
ings of preparations in my own possession. 

R. E. DINWIDDIB. 

FATETTEVILLE, ARK., July 19, 1894. 



TABLE OP CONTENTS. 



PART FIRST. 
Generalities upon Microbes. 

I. Microbes in the Static Condition. 

Definition 11 

Forms 11 

Organization and chemical composition 14 

Occurrence and distribution in nature *. 14 

II. Physiology of Microbes. 

Foods of microbes 19 

Digestion of microbes 19 

Eespiration of microbes 21 

Nutrition of microbes ' 22 

Movements of microbes 24 

Generation, multiplication 25 

Action of the media on microbes 27 

Action of microbes on the media 30 

Role of bacteria in nature 30 

Fermentations 81 

Putrefaction 32 

Role of bacteria in the normal organism 33 

Digestive action of microbes , 33 

Putrefaction of cadavers . 34 

R61e of microbes in the organism in the pathological con- 
dition 36 

Classification ... 37 

PART SECOND. 
Generalities upon Pathogenic Microbes. 

I. Pathogenic Microbes in the Static Condition. 

Saprogenic or eaprophytic germs , 39 

(v) 



vi Contents. 

Pathogenic germs 40 

Conditions of existence of pathogenic germs in external 

media and in the economy. Sources of infection 40 

Contagious obligatory parasitic microbes 40 

Contagious facultative microbes 41 

Non-contagious facultative microbes , 42 

Distribution of pathogenic germs 43 

Modes of contagion 52 

Immediate contagion 52 

Direct contact 53 

Heredity 53 

Mediate contagion 54 

Absorption of pathogenic microbes 55 

El. Physiology of Pathogenic Microbes. 

Action of microbes upon the organism 61 

Pathogeny of the local changes 61 

Pathogeny of general and remote changes 63 

Receptivity 65 

Immunity 70 

Reaction of the organism against microbes 74 

Phagocytosis 74 

Bactericidal state 76 

Elimination of microbes 78 

Modifications of virulence , . 79 

Evolution of the bacterial disease 80 

Incubation 81 

Latent microbism 81 

Specificity of pathogenic microbes 82 

III. Transformation and Destruction of Pathogenic Microbes 

in their Relation to Hygiene and Therapeutics. 
Morphological and physiological variations of pathogenic 

microbes 85 

Attenuation 87 

Preventive inoculations ; vaccinations 94 

Destruction of pathogenic microbes 97 

IV. Methods of Determination of Pathogenic Microbes. 

Basic and acid colors 104 

Examination of liquids 106 

Examination of organic pulps 107 

Examination of sections 107 



Contents. vii 

Mounting of preparations 108 

Single stains 109 

Double stains 110 

Method of Loffler.. . . 110 

Method of Malassez and Vignal 110 

Method of Gram , 110, 111 

Method of Weigert Ill 

Method of Kiihne 112, 113 

Method of Berlioz . . 114 

Method of staining spores 115 

Culture of germs 115 

Sterilization .115 

Culture media 119 

Isolation of bacteria 129 

Inoculation of culture media 130 

Culture ovens 132 

Experimental contagions 139 

PART THIRD. 
Microbic Diseases Individually Considered. 

Microbic diseases consecutive to wounds 145 

Suppuration 147 

Pyaemia 152 

Septicaemia 153 

Pasteur's septicaemia 160 

Septicaemias of the rabbit 167 

Koch's experimental septicaemia 167 

Spontaneous septicaemias of the rabbit 168 

Hemorrhagic septicaemias 323 

Chicken cholera 170 

Infectious enteritis of chickens 177 

Epizootic dysentery of chickens and ducks 177 

Duck cholera 178 

Bateridian charbon /. 180 

Symptomatic charbon 1 94 

Kouget of the pig 203 

Pneumo-enteritis or cholera of the pig 208 

Pneumo-enteritis of the sheep 216 

Infectious, pneumonia of the pig 218 

Tuberculosis.. . 221 



viii Contents. 

Tuberculosis, diagnosis of doubtful cases 233 

Tuberculosis and scrofula 246 

Tuberculosis of mammals and tuberculosis of fowls 247 

Tuberculosis, zoogloeic 251 

Tuberculosis, bacillar of Courmont 253 

Tuberculin 235 

Glanders 254 

Glanders, diagnosis of doubtful cases 260 

Mallein 260 

Epizootic lymphangitis 265 

Strangles 265 

Contagious acne of the horse 267 

Actinomycosis 268 

Botryomycosis 280 

Bovine farcy 282 

Tetanus . . 284 

Diphtheria .295 

Rabies 299 

Equine typhoid fever 311 

Contagious pneumonias of the horse 313 

Contagious pleuro-pneumonia of cattle 316 

Septic pleuro-pneumonia of calves 320 

Epizootic abortion . , ' 324 

Contagious mammitis of milch cows 326 

Gangrenous mammitis of milch ewes 329 

Diseases of milk 329 

Bacterial hsemoglobinuria of cattle 332 

Distemper of young dogs 334 

Phosphorescent meats 338 

Appendix 340 



INTRODUCTION. 



"Works which treat of Microbiology are quite nu- 
merous, but none offer a concise and complete expo- 
sition of the accepted facts on the subject, the appli- 
cation of which is within the reach of students and 
practitioners. Such outline we would give here in 
the hope that both may -be benefited thereby. 

It would seem that this publication has some pros- 
pect of being well received. Besides the fact that 
the veterinarian in daily practice is under the neces- 
sity of having recourse to the teachings of microbi- 
ology, the inspection of meat and the supervision of 
the sanitary police duties which have devolved upon 
him make it his imperative duty to neglect no 
means of diagnosis which science places at his dis- 
posal. 

"We do not mean to assert that the diagnosis of in- 
fectious diseases necessitates in all cases a search for 
the pathogenic microbes, but, recognizing the im- 
portance of the pathological anatomy and clinical 
symptoms, we believe that the demonstration of 
these germs is of much higher value. We will even 
say that, in unfortunately too many cases, the recog- 



x Introduction. 

nition of the germs is the only mode of definitely 
establishing the nature of a lesion. 

Every one can understand how desirable it is that 
the practitioner, meat inspector or sanitary veterina- 
rian, whose decisions very frequently run counter to 
some particular interest, should pronounce himself 
only after having made use of this last resource 
which will protect him from scientific mistakes a.nd 
contradictions, always much to be regretted. 

A like exactitude is to be desired in private prac- 
tice, where it will form a basis for sound therapeutics. 

We will briefly trace the history of those microbes 
which may be of interest to the practitioner, and de- 
scribe the technique of those investigations which he 
may daily be called upon to make. Our study will 
embrace three divisions. In the first we will briefly 
consider the subject of microbes in general ; in the 
second we will study pathogenic germs collectively ; 
and in the third notice those particular microbes 
which occasion disease in animals and even in man- 
kind. 



OF VETERINARY MICROBIOLOGY. 



PART FIRST. 

MICROBES CONSIDERED IN GENERAL. 

Microbes may be considered successively in the 
static and in the functional or physiological condition. 



CHAPTER I. 

MICROBES IN THE STATIC CONDITION. 

Definition 1. Forms of microbes ; 2. Organization and chemical 
composition ; 3. Situation and distribution. 

The names Microbes, Bacteria, Vibrios, Schizomycetes, 
Schizophytes, have been given to unicellular micro- 
scopic beings placed at the bottom of the scale of 
the vegetable kingdom. These beings, destitute of 
chlorophyll, live at the expense of complex organic 
substances, which they reduce -to the condition of 
simple mineral compounds. 

I. Forms of microbes. 

1. Typical forms. The form of microbes is that of 
a rounded corpuscle or of a rod. The latter may be 
straight, undulated, or spiral. 



12 



Manual of Veterinary Microbiology. 



Fig. I. 




Rounded microbes or cocci have received the follow- 
ing names, according to the manner in which they are 
grouped: 

Micrococcus: cocci isolated, 

. 1,1); 

Diplococcus: cocci arranged 
in pairs, (2) ; 

Streptococcus: cocci ar- 
ranged in linear series, in 
chains, (3); 

Micrococcus tetragenus: cocci 
arranged in groups of four, 

(4); ' 

Sarcina : cocci arranged in 
tetrads so as collectively to 
form a cuhe. 

Staphylococcus : cocci associated in clusters ; 

Zoogloea: cocci associated in large numbers in an 
amorphous matrix, (5) ; 

Ascococcus : cocci associated in large numbers in an 
amorphous matrix and inclosed in an enveloping mem- 
brane, (6). 

The elongated microbes are cylindrical or fusiform 
rods, or have the shape of a bell clapper. They have 
received the following names : 

Bacillus : rods short and straight, (7) ; 

Leptothrix : rods long and undulating, (8) ; 

Cladothrix: rods long, straight, and branching, (9). 

Spiral microbes are in the form of an arc of a circle, 
or are spiral. They appear, however, with rectilineal 
forms when their curvature is directed toward the ob- 
jective. The following terms are used to designate 
them : 



Microbes in the Static Condition. 13 

Vibrio : microbes spiral, short, (10) ; 

Spirillum : microbes spiral, long and rigid, (11) ; 

Spirochcete: microbes spiral, long and flexible, (12). 

2. Forms of involution, of degeneration. Under spe- 
cial conditions unfavorable to their nutrition bacteria 
may assume abnormal aspects, such as swelling in the 
form of a club, at which place the protoplasm, becomes 
clearer : these special forms are called forms of invo- 
lution, (14). They have been established in the acti- 
nomyces, the bacillus of Koch, etc. 

3. Polymorphism. Microbes are essentially poly- 
morphic. Recent researches have shown that the 
same microbe may assume very different aspects, ac- 
cording to the medium in which it lives. Thus, the 
germ of the pyocyanic disease presents itself succes- 
sively as a bacillus, a spirillum, and a micrococcus. 
The bacillus of Pasteur's septicemia grows in long 
filaments in the blood, in short bacilli in the subcu- 
taneous cellular tissue. The bacillus of symptomatic 
charbon, cultivated in bouillon containing glycerin 
and sulfate of iron, takes the form of a clove. With 
the same germ, therefore, we can obtain several mor- 
phologically distinct individualities. 

The dimensions of microbes are as variable as their 
form ; in all cases these are expressed by a few thou- 
sandths or even fractions of the thousandth of a milli- 
meter.* 

* [The dimensions of microscopic objects are usually expressed 
in Microns. A micron is the one thousandth part of a millimeter, 
and is designated by the Greek letter p. The dimensions of 
microbes, expressed in the original of this work in decimal frac- 
tions of a millimeter, have been rendered in the translation as 
microns. Thus Omm., 005=5/u. D.] 



14 Manual of Veterinary Microbiology. 

II. Organization of microbes. 

Structure and chemical composition. The structure 
of microbes may be compared to that of a cell with- 
out nucleus. The existence of the latter is not gener- 
ally admitted, although some authorities believe they 
have observed such a structure. 

The content is a kind of protoplasm which Nenki 
has designated by the name of mycoprotein, a homo- 
geneous, or sometimes granular, substance generally 
devoid of color. Under certain circumstances it may 
contain brilliant corpuscles (spores), starch grains (in 
the Sarcina), and granules of sulphur (Beggiotoa). 

The periphery is formed by a thin and flexible, or 
thick membrane, the nature of which is not well 
known. Most authors look upon it as a carbo-hydrate 
allied to cellulose; its resistance to acids and alkalies 
seems to support this view. Others regard it as a 
layer of mycoprotein differentiated from the proto- 
plasm. 

Sometimes this membrane emits vibratile cilia ; at 
other times it is surrounded by a zone of a mucilagi- 
nous aspect, capable of swelling up in water and form- 
ing a transparent capsule to the germ (pneumococcus). 

III. Situation and distribution of germs in nature. 

The part which these beings play in nature being 
known, it is easy to understand that they should be 
found wherever there is organic matter to be reduced. 
"We find them, indeed, pullulating in all places where 
there are no special conditions prejudicial to their life. 
"We will notice their distribution in the most impor- 



Microbes in the Static Condition. 15 

tant media : atmospheric air, waters, soil, foods, the 
living organism, dwellings, vehicles, clothing, etc. 

Air. The germs which are found suspended in the 
atmosphere can not multiply there, and hence come 
from other media from which they are carried off by 
atmospheric currents along with pulverulent matters. 
They are deposited in the calm (tranquil air, hollow 
places). Their number and nature vary with climatic 
and other conditions ; their number increases during 
desiccation of the soil (summer), and diminishes after 
rains. They are most numerous in inhabited places 
and in the vicinity of marshes, whilst the air of moun- 
tains and of the surface of seas is almost completely 
free from them. 

The germs of the air do not long resist the com- 
bined action of oxygen and light ; nevertheless, through 
the agency of the winds their effects may be mani- 
fested at great distances. 

The study of atmospheric germs is made by simple 
enumeration with the microscope or by various cul- 
ture processes. The latter method is much to be pre- 
ferred since it allows of the separation of dead germs, 
the number of which is very large. 

Waters. Subterranean waters, having filtered 
through thick layers of earth, are free from all germs, 
but quickly become infected on contact with the surface 
soil and the air. The waters of wells are always in- 
fected ; their pollution is, moreover, easy to under- 
stand : the masonry having no support at the bottom 
eventually sinks down and fissures are produced 
through which infiltrate the waters of the neighboring 
surface soils often impregnated with germs. For this 



16 Manual of Veterinary Microbiology. 

reason wells should not be built in proximity to cis- 
terns, cess-pools or dung-hills. 

The deep waters best protected against infection 
are those of artesian wells. 

Surface waters are always very rich in germs, the 
nature of which is extremely variable ; the majority 
are ubiquitous germs, pathogenic being much less fre- 
quent. Stagnant waters especially favor the multipli- 
cation of germs. 

Soil. Whilst the rocks and the virgin soil from the 
depths are free from all germs, these occur in large 
numbers in the superficial layers. Their number and 
nature vary infinitely according to location, season, 
winds, the physical constitution and chemical com- 
position of the soils, etc. Their multiplication grad- 
ually decreases as the depth increases. Water, in 
filtering through the ground, yields up the germs 
which it contains as well as the soluble matters which 
serve for their nutrition. 

It may be said, in truth, that germs in way of pul- 
lulation, through their power of penetrating the cap- 
illary spaces, should of themselves sink below the 
layer of soil in which the waters deposit them. This 
vegetation, however, is itself impeded by unfavorable 
conditions of temperature and nutrition, the absence 
of oxygen, etc. Hence in good filtering soil germs 
are no longer found at a depth of three meters. 

Foods. Vegetable foods (fodders, oats, etc.) are al- 
ways contaminated with germs derived from the air, 
the soil, or waters. Animal foods are generally con- 
taminated by contact with the air. Foods, whether 
of vegetable or animal origin, are especially favorable 
to the multiplication of microbes. The various means 



Microbes in the Static Condition. 17 

employed for the preservation of foods have no other 
aim than to protect them against the invasion or the 
destructive action of these organisms. When these 
means are defective or powerless to arrest the evolu- 
tion of the germs which have been deposited there, 
various changes supervene which diminish the in- 
trinsic nutritive value of the foods and may even ren- 
der them detrimental to the health of man or of ani- 
mals (damaged hay, putrid meat). The contamina- 
tion of foods by pathogenic germs, properly so called, 
will be studied later. 

Houses and vehicles. The walls, floor, and ceiling, as 
well as the mangers and racks, of houses occupied by 
animals are constantly liable to receive the germs 
which are borne in the atmospheric dust, cleansing 
waters, solid dejections, litter, foods, etc. 

Vehicles (wagons, etc.) serving for the transport of 
animals may be contaminated by microbes in the same 
way as houses. 

Harness, blankets, tools, and other objects. It is easy 
to understand that these objects will most frequently 
be contaminated either by the various methods men- 
tioned above or by their contact with the animal for 
the use of which they are destined. 

Organism. After what we have said of the nutrition 
of germs, we may expect to encounter them in all 
parts of the economy which are in direct relation 
with the air, or with solid and liquid media. 

The digestive canal throughout all its course con- 
tains in large numbers various microbic species which 
have been -carried there by food and drink. In the 
mouth we find especially a leptothrix, spirochsete, and 
2 



18 Manual of Veterinary Microbiology. 

vibrios; from it have also been isolated pathogenic 
germs staphylococcus pyogenes, etc. 

The stomach contains especially sarcinae, yeasts, 
and elongated bacteria, whilst the intestines contain 
large numbers of bacilli. Micrococci and elongated 
non-sporulated bacteria are killed by contact with the 
gastric juice, and, consequently, do not multiply in 
the intestine. 

The mucus of the anterior respiratory passage is 
likewise always contaminated with germs which have 
been deposited by the inspired air, the latter itself be- 
ing thus punned so as to emerge free from all germs. 
Hence, it apppears that disease infection can not take 
place through the expired air. The ocular mucosa 
and genito-urinary mucosa near the external openings 
contain also a certain number of microbes. 

Finally, these are found lodged upon the skin, the 
perspiration and sebaceous secretion along with the 
epidermic debris -normally cast off constituting a good 
medium for their preservation. 

The blood of healthy animals is free from germs. 
In the pathological condition, on the other hand, most 
of the tissues and fluids of the organism may become 
the seat of the evolution of bacteria. 



Physiology of Microbes. 19 



CHAPTER II. 

PHYSIOLOGY OP MICROBES.' 

1. Digestion. 2. Respiration. 3. Nutrition. 4. Movements. 
5. Generation and multiplication. 6. Action of the media 
upon microbes. 7. Action of microbes upon the media. 
8. Classification. 

I. Digestion. 

1. Foods. Microbes being destitute of chlorophyll 
require for their nutrition organic products already 
formed; consequently they must nourish themselves 
at the expense of vegetable or animal substances. 
They borrow nitrogen from albuminoid .substances or 
their derivatives as well as from ammoniacal salts, and 
occasionally, in part, from nitrates ; carbon and hy- 
drogen from hydrated carbonaceous substances 
sugar, glycerin, and salts of malic, tartaric and acetic 
acids. 

They require also mineral substances sulfates and 
phosphates of sodium, potassium, and magnesium. 

They are very sensitive to the chemical composition 
of the nutrient medium in which they live ; traces of 
certain substances, as well as the absence of others, 
can bring about profound alterations in the manifesta- 
tions of their vitality. 

Their medium should be slightly alkaline and very 
aqueous. Excess of acidity or of alkalinity, acidity 
especially, is prejudicial to their growth. 

2. Digestion. The foods of microbes, like those of 



20 Manual of Veterinary Microbiology. 

animals and vegetables, independent of the chloro- 
phyllic function of the latter, require to undergo cer- 
tain modifications preparatory to assimilation. These 
modifications, representing the digestion of microbes, 
consist of a hydration accompanied or not by a split- 
ting up of molecules. 

This phenomenon is accomplished by means of sol- 
uble ferments secreted by the germs, and it is remark- 
able that these digestive ferments are the same as 
those which are found in higher beings ; thus, in the 
case of microbes as for these last, starch is transformed 
into dextrin by a diastase^ 1 ) called amylase correspond- 
ing to vegetable diastase, ptyalin, and to the amyla- 
ceous ferment of the pancreatic juice. Cane sugar is 
split up into glucose and levulose by a diastase or 
sacrase identical with the invertin of the beet-root 
and with the inverting ferment of tne intestinal juice ; 
albuminoid substances are peptonized by microbes 
through the secretion by the latter of a special pepsin 
or casease ; in the case of the casein of milk the ac- 
tion of this last substance is preceded by that of a dias- 
tase analogous to rennet which, like the latter, deter- 
mines the coagulation of the milk. 

When we consider the great number of germs con- 
tained within the digestive canal, the comparison 
which we have just made between digestion in mi- 
crobes and in the higher beings, indicates the possi- 
bility of an adjuvant action of the former in the di- 
gestion of animals. 

0)The name diastase, formerly limited to the amylaceous fer- 
ment of vegetables, is now synonymous with soluble ferment, 
thus amylase, sucrase, pepsin, rennet (pr^sure), are diastases^or 
xymases. 



Physiology of Microbes. 21 

These are not the only diastases secreted by mi- 
crobes. We are far from knowing all of them ; they 
vary, naturally, according to the special nature of 
their food. We have noted the principal of them in 
order to bring out the general mode of the nutritive 
process in microbes. 

II. Respiration. 

The study of the respiration of microbes is of great 
interest. Obviously, all require oxygen which, in ox- 
idizing alimentary substances, supply the heat neces- 
sary for the maintenance of life, for multiplication 
and motion, etc. Many of them borrow it in the free 
state from the atmosphere or from water (aerobes), but 
there are others which appear incapable of enduring 
free oxygen, hence require to live protected from the 
air (anaerobes). These last act upon certain organic 
substances by a sort of internal combustion ; they re- 
duce these substances into carbonic acid and into other 
molecules generally less complex, but still susceptible 
of oxidation, setting free a certain amount of energy, 
as the aerobic germs on their part do by a true com- 
bustion. 

One of the prominent characters of anaerobic germs 
when they are nourished at the expense of quaternary 
substances consists in the disengagement of abundant 
gaseous products, among which we find, besides 
carbonic acid, nitrogen, ammonia, and ammoniacal 
compounds (trimethylamin, etc.), sulfuretted and 
phosphoretted hydrogen, etc.; these mixtures emit a 
peculiarly fetid odor (putrid gases). When they es- 
pecially reduce ternary products these anaerobes give 
as gases carbonic acid, hydrogen, and hydro-carbons. 



22 Manual of Veterinary Microbiology. 

A certain number of germs accommodate themselves 
equally well to both these ways of life; in the air 
they are aerobic, in its absence they become anaerobic. 
This double faculty has been expressed by the term 
aero-anaerobic. 

III. -Nutrition. 

1. Absorption and assimilation. Absorption of food 
takes place by osmosis. A part of the principles ab- 
sorbed is utilized for the elaboration of plastic mate- 
Hal ; another part behaves as a respiratory food. The 
formation of plastic material must be considerable 
when we take into account the excessively rapid mul- 
tiplication of micro-germs. 

The respiratory foods serve especially for the pro- 
duction of the different forms of work performed by 
the elements, and which are represented by the phe- 
nomena of assimilation, growth, locomotion, heat, 
sometimes light (phosphorescence of meat and fish). 

Their intimate nutrition is little known. Chemi- 
cally, the point of departure of the nutritive action is 
quite different according to the case. Some require 
albuminoids, whilst others draw their nitrogen from 
azotized products with molecules of much less com- 
plexity: leucin, tyrosin, xanthin, etc.; others again 
borrow it from trimethylamin and from ammoniacal 
salts. The same variety is observed in the case of 
non-nitrogenous foods. 

This peculiarity accounts for the successive appear- 
ance of different bacteria in an organic medium aban- 
doned to the external air. As this medium becomes 
more and more exhausted those species of germs suc- 
cessively appear whose lesser requirements permit of 



Physiology of Microbes. 23 

their living at the expense of the nutritive residue of 
those which have preceded them. 

2. Disassimilation^ excretions, secretions. From the 
preceding considerations it results that disassimilation 
in microbes ought to give very varied residues. These 
residues naturally depend upon the food, upon the 
species of germ, and upon the special conditions in 
which their evolution is accomplished (temperature, 
aerobic or anaerobic nature, etc.). 

Of these residual or excrementitial products of mi- 
crobes some are gaseous (carbonic acid, hydrogen, car- 
buretted hydrogen, sulfuretted hydrogen, ammonia), 
some volatile (trimethylamin, alcohol, formic, acetic, 
butyric acids, etc.), some fixed (lactic and malic acids, 
leucin, taurin, tyrosin, etc., etc.). 

The nutrition of germs may give rise to coloring 
matters, such germs being called chromogenic. The 
coloring matter thus produced may be soluble or in- 
soluble; in the former case it diffuses in the fluid 
media in which the germs occur, an instance of which 
may be seen in the germ of blue milk. 

Among the number of the substances resulting 
from the nutrition of microbes we have to mention 
the ptomaines. 

Ptomaines are ammoniacal compounds acting the 
part of bases, and which, upon the higher beings, 
have often effects analogous to those of the vegetable 
alkaloids, which they resemble in every respect. 

In a general way the residual products are noxious 
to the germs from which they spring : 0.8 per cent of 
free butyric acid arrests the butyric fermentation of 
lactate of lime. 

The diastases must also be cited among the products 



24 Manual of Veterinary Microbiology. 

of the nutrition of microbes ; they are secreted for 
the requirements of digestion, as we have already 
seen. 

Later, we shall have occasion to see, when consid- 
ering the role of microbes, that their nutrition is the 
determining cause of the chemical reactions which 
characterize fermentations and putrefaction. 

IY. Movements of microbes. 

Some bacteria are immobile (the majority of round 
and some elongated bacteria), others are gifted with 
the faculty of moving themselves in the fluids in 
which they live. 

The kind of movement varies with the species con- 
cerned ; sometimes the element, maintaining its recti- 
lineal direction, performs a simple, more or less regular 
oscillation around an imaginary longitudinal axis ; at 
other times it undergoes a slight inflection in the di- 
rection of its length and straightens itself again alter- 
nately ; at other times, again, it assumes a flexuous ap- 
pearance simulating the movements of a snake ; some 
even wind themselves around in corkscrew fashion. 

The motion of a certain number of bacteria is de- 
pendent upon the presence of vibratile prolongations, in 
others these movements seem to depend upon contrac- 
tions taking place within the body of the element. 

In all they are directly dependent upon nutrition 
the integrity of which is necessary to their produc- 
tion. 

Light and the fluidity of the media are conditions 
which favor them. 



Philosophy of Microbes. 25 

Y. Generation, multiplication. 

1. Spontaneity. Formerly it was supposed that mi- 
crobes originated by spontaneous generation in putre- 
factive media; this origin, indeed, was accepted for 
all beings the mode of reproduction of which was un- 
known. The progress of the natural sciences first 
considerably restricted the scope of this theory, which 
the experiments of Pasteur triumphantly and abso- 
lutely combatted. Although it can not be denied that 
at a period in the remote past organized matter must 
have been formed spontaneously at the expense of 
mineral matters, it seems well established now that 
the molecular association which tends to the constitu- 
tion of protoplasm is no more produced, at least 
within the conditions of observation accessible to man, 
except at the expense of a pre-existent being. We 
therefore have to consider here, from a practical 
point of view, only the reproduction of germs by mul- 
ti plication. 

2. Fission. Microbes multiply principally by fission. 
The cells of which they are composed become elon- 
gated, then divided into two by a transverse groove ; 
the two segments which result from this division may 
separate and live independently or may remain united 
so as to form agglomerations of various kinds ; for 
example : the chains or chaplets of micrococci 
which adhere end to end ; the zoogloea to which the 
same micrococci give rise when united in mass by a 
gelatinous substance, the jointed filaments of the an- 
thrax bacillus, etc. 

Fission usually takes place in one direction only, 
' 3 



26 Manual of Veterinary Microbiology. 

but there are .bacteria in which the division takes 
place in two crossed directions (micrococcus tetra- 
genus), or even in three directions (sarcina) ; in this 
last case the bacteria, the secondary elements of which 
remain united, take the form of a cube. 

3. Sporulation. Multiplication by fission appears to 
be the only mode possessed by microbes of spherical 
form ; in the majority of others we recognize a second 
mode, sporulation (13). This consists in the forma- 
tion within the bacteria of brilliant points which, are 
apparently the result of a condensation of the orig- 
inal protoplasm, whilst the latter at the same time be- 
comes very clear. These brilliant points are the 
spores ; they are set at liberty by the destruction of 
the cell which has produced them and when they 
find themselves in good conditions of temperature and 
humidity, and in a suitable medium, they reproduce 
the bacteria as, in the higher forms of vegetation, the 
seed gives origin to the entire plant. 

Spores show a remarkable resistance to the action 
of the common causes of destruction of microbes. 
They almost never develop in the media in which they 
have taken birth. 

Botanists recognize, besides fission, which for them 
is only a form of growth, or vegetation, two methods 
of sporulation or fructification. The first and best 
known is endosporulation which we have just described; 
the second which it is not always easy to distinguish 
from fission has received the name arthrosporulation. 
It is characterized by the production by fission, at the 
expense of cells performing the function of repro- 
ductive elements, of new cells which differ from those 
obtained by ordinary fission by their state of latent 



Philosophy of Microbes. 27 

life and by the thickness and resistance of their en- 
veloping membrane. They are . generally larger than 
the spores begotten by endosporulation and resemble 
cysts. It is this distinction in the mode of formation 
of spores which serves as the fundamental basis of 
the classification of Ghiignard which we will reproduce 
later. 

It is not without interest to have an idea of the 
power of multiplication possessed by these micro- 
organisms. If we regard a bacterium as dividing 
itself into two after one hour, we will have four of 
them in two hours, and in twenty-four hours, 16,000,- 
000. After forty-eight hours we will have the fabu- 
lous number of 280 trillions. We can estimate from 
this the ravages that germs introduced into the blood 
must produce when they find there conditions favor- 
able to their pullulation. 

VI. Action of the media upon microbes. 

The medium has necessarily a great influence upon 
microbes. Besides supplying them with food it is 
capable of modifying their vitality in different de- 
grees. 

Certain agents or conditions have the power of 
bringing bacteria to the condition of latent life, of 
changing their usual mode of activity, or even of de- 
stroying them. These agents or conditions are of a 
nature either mechanical, physical, chemical, or phys- 
iological. 

1. Mechanical influences. These are badly deter- 
mined; some authors claim to have observed that 
oscillations impressed upon cultures of microbes are 



28 Manual of Veterinary Microbiology. 

adverse to their multiplication ; other experimenters 
have arrived at directly opposite results. 

2. Physical influences. a) Humidity. Water is in- 
dispensable to microbes; a medium containing less 
than sixty per cent of this liquid arrests their multi- 
plication. Desiccation causes all "active life to cease 
(latent life) and may in this way finally lead to their 
death. 

Similarly, it is to their deficiency in water content 
that we must attribute the preservation of vegetable 
juices by sugar, of meat pastry by fat, etc. 

6) Temperature. A temperature too high or too 
low is detrimental to microbes. They are generally 
more sensitive to an elevation of temperature than to 
a depression. The vegetative forms are killed by two 
hours' exposure to a temperature of 48 to 60 C. The 
spores, however, are much more resistant and require 
for their destruction a temperature of 140 when they 
are in a dry medium, 100 when they are moist. 

The temperature of predilection of microbes is from 
20 to 39 ; above the latter temperature disturbances 
of microbic activity, either temporary or permanent, 
are liable to ensue (principle of attenuation by heat). 

Cold arrests the multiplication of microbes (latent 
life), but kills them only with difficulty. Some have 
been exposed to a temperature of 105 without their 
vitality being completely destroyed. 

c) Light. Light is a puissant cause of destruction 
to microbes ; it excites oxidation of their constituent 
principles and especially of the hydro-carbonaceous 
substances ; its action is quite rapid and continues even 
after the germs are again removed from the light. 

d) Electricity. -The action of continuous and of 



Physiology of Microbes. 29 

induction currents have been studied. This action, 
however, is but little known ; in the case of continu- 
ous currents it is necessary to take into account the 
electrolysis which separates the acids from the bases 
and transfers these new molecules to the two poles. 
The acid reaction of the positive pole is opposed to 
microbic pullulation at this point; the alkalinity of 
the negative pole is less energetic in its action. The 
current may act upon the germ itself and interfere 
with its multiplication, especially when strong cur- 
rents are employed. It is not inadmissible to hope 
that, the influence of electricity becoming better 
known, its effects may some day be utilized in the 
treatment of infectious diseases. 

3. Chemical influences. The exhaustion of the nu- 
tritive medium, accumulation of residual products, 
excess of alkalinity and more especially of acidity, 
oppose and may even arrest the multiplication of 
germs. 

Certain chemical substances exert toxic effects upon 
microbes, this toxicity varying with the microbic 
species concerned. The rational application of this 
action constitutes the basis of antisepsis from which 
hygiene and medicine have already derived great 
profit. 

4. Physiological influences. When several microbic 
species occur in the same place they may oppose each 
other and then a veritable struggle for existence en- 
sues. From this concurrence may result the annihila- 
tion of the least favored species; the medium, the 
number of germs, and various other circumstances 
may intervene here. In putrefactive media in which 



30 Manual of Veterinary Microbiology. 

the bacterium termo and bacillus subtilis live we do 
not find the bacillus septicus. 

Under other circumstances it may happen that the 
two associated germs assist each other, the one pre- 
paring alimentary materials for the other, or, it may 
be, protecting the other from the noxious action of 
certain agents (association of aerobic and anaerobic 
germs). 

We ought to mention in this connection the influ- 
ence exerted upon microbes by the tissue elements of 
animals in which they sometimes live (phagocytosis) ; 
we will enter more into the details of this subject in 
the second part. 

VII. Action of microbes upon the media. 

1. R6LE OF THE BACTERIA IN NATURE. 

From a general biological point of view the r6le of 
microbes consists in reducing to the condition of sim- 
ple inorganic compounds the organic matter built up 
by vegetables and incorporated by animals. The dis- 
assimilation which occurs in these beings correlative 
with nutrition destroys a part of this organic matter 
and reduces it to the condition of carbonic acid, 
water, and salt's. This constantly occurring decom- 
position, however, not only fails to completely min- 
eralize the substance upon which it acts, but, in addi- 
tion, at the death of animals and plants an amount 
of elaborated substance remains which, in future, is 
exempt from this cause of destruction. It is here 
that microbes or, more accurately, ferments enter on 
the scene. 

" The life of the larger forms of vegetation builds' 



Physiology of Microbes. 31 

up in nature at the expense of the solar heat, sub- 
stances whose production requires a certain expendi- 
ture of force. It is in these endothermic subtances 
that the lower organisms implant themselves. From 
the energy which they find there stored up they bor- 
row a portion for the construction of their own tis- 
sues, which renders them up to a certain point inde- 
pendent of external conditions. Another portion is 
used to convert into the gaseous condition substances 
originally fluid or solid. Another, finally, is trans- 
formed into sensible heat and serves to elevate the 
temperature of the fluid in which all these phenomena 
occur, and, as a consequence, to accelerate their pro- 
duction." (Duclaux.) 

To accomplish this immense work the ferments are 
endowed with an intense destructive power, and ope- 
rate, thanks to the rapidity of their multiplication, in 
innumerable legions. 

We must here briefly refer to fermentations and 
putrefaction. 

Fermentations. Fermentations are always the result 
of the intervention of micro-organisms. They consist 
in modifications of special organic substances tending 
to the formation of simpler products in which the heat 
of total combustion is less than that of the ferment- 
able substances from which they are derived. The 
difference between these quantities of heat represents 
the amount of energy appropriated by the germ for 
its nutritive requirements, and the reaction by which 
the fermentation is characterized has no other object 
than the liberation of this energy. 

Fermentable substances are comparatively few in 
number; they are usually bodies rich in oxygen 



32 Manual of Veterinary Microbiology. 

carbo-hydrates, polyatomic alcohols, the lower fatty 
acids, and albuminoids. According to the substance 
predominating in their products, fermentations are de- 
scribed as alcoholic, acetic, lactic, butyric, viscous, 
ammoniacal, etc. 

Microbes capable of developing fermentations are 
called zymogenic. 

Putrefaction. By putrefaction is meant the decom- 
position of the substance of organized beings through 
the agency of microbes. This decomposition super- 
venes shortly after death ; exceptionally it is observed 
during life as we will see in the special part of this 
work. The microbes which determine it are called 
septogenic. It consists in a series of fermentations so 
associated that the products of the one form the point 
of departure for the following. These fermentations 
occur simultaneously and take effect upon the various 
immediate principles of the organism: albuminoid 
substances, hydrocarbonaceous bodies, etc. 

But the decomposition of these last named princi- 
ples giving rise to fermentations simpler and better 
known, and specially denominated as such, the term 
putrefaction refers more particularly to the microbic 
degradation of albuminoid molecules. 

Many microbic species are concerned in putrefac- 
tion, these species varying according to the case ; there 
are some which are quite frequently met with, such 
as bacterium termo, bacillus subtilis (hay bacillus), and 
bacillus septicus, micro-germs whose characteristics will 
be indicated later, we always meet with an association 
of aerobic and anaerobic organisms. 

The bodies which arise during putrefaction are nu- 
merous : hydrogen, sulfuretted, carburetted and phos- 



Physiology of Microbes. 33 

phoretted hydrogen ; ammonia, carbonic, formic, acetic, 
butyric, and lactic acids, etc.; amines, trimethylamin, 
propylamin, etc.; indol, scatol, tyrosin, ptomaines, etc. 
The term saprogenic is applied more particularly to 
those microbes which excite putrefaction with disen- 
gagement of a peculiarly fetid odor. 

2. R6LE OP BACTERIA IN THE NORMAL ORGANISM. 

In a general way the microbes which live within or 
upon healthy individuals behave as commensals with- 
out giving rise to the slightest disturbance in their 
host. 

Nevertheless, as there are almost always, among the 
germs distributed on the normal organism, a certain 
number which are pathogenic, we can understand that 
their presence constitutes a permanent source of dan- 
ger for the economy. This danger exists upon all 
contaminated surfaces the germs being able to invade 
the organism when an accidental abrasion occurs. 

Digestive action. The germs lodged within the di- 
gestive canal have a more interesting r6le, for they seem 
to place themselves at the service of their host in 
order to assist in the preparation of his foods, in his 
digestion. Bienstock has isolated from faeces a 'bacil- 
lus which converts albumen into peptones ; the bacil- 
lus amylobacter, which is found in the stomach of 
ruminants, secretes a soluble ferment which acts upon 
starch and cellulose. 

M. Abelous has collected from the stomach sixteen 
species of microbes whose resistance to the action of 
the gastric juice he has verified. The study of their 
digestive action gave very interesting results. Albu- 
men, casein, fibrin and gluten were rapidly and com- 



34 Manual of Veterinary Microbiology. 

pletely peptonized by several of them. Cane sugar 
was inverted by eight different species. Eleven species 
more or less completely converted starch into sugar. 
The conjoined action of these different micro-organ- 
isms upon a complex food must be considerable. 

Putrefaction of dead bodies. .-The healthy intestinal 
mucosa forms an effective barrier to the invasion of 
germs pullulating within the intestine; after death the 
cells having lost their power of resistance are rapidly 
dissolved by the diastases which these microbes secrete, 
and the latter penetrate within the tissues. They are 
first found in the peritoneum and on the surface of the 
abdominal viscera ; they multiply in the blood of the 
mesenteric veins and extend along the portal vein, 
from which they progress toward the heart ; thus, in 
various ways, they more or less rapidly invade the 
whole economy. 

These anaerobic germs find in the organism deprived 
of oxygenated blood the most favorable conditions for 
their multiplication. Hence putrefaction is the more 
rapid in proportion as the blood is poorer in oxygen 
at the time of death, for example, in animals dead 
from charbon. In some cases the lack of oxygen is 
seconded by the absence of coagulation of the blood 
and of cadaveric rigidity, conditions which, by main- 
taining the fluidity of the medium, render microbic 
invasion more easy. 

Putrefaction of cadavers is, therefore, primarily the 
effect of anaerobic germs coming from the intestinal 
surface; the anaerobes of other surfaces are in fact 
paralyzed in most cases by contact with the oxygen 
of the atmosphere. Putrefaction differs somewhat ac- 
cording to the surroundings in which it occurs. 



Physiology of Microbes. 35 

Putrefaction in the air. The multiple fermentations 
developed by microbes cause softening of the paren- 
chyma, dissolution of the blood globules, infiltration 
of coloring matters in dependent parts, gaseous disen- 
gagements which permeate the connective tissue and 
elevate the skin, bloating, swelling of various parts of 
the body, rupture of the surfaces and oozing of sani- 
ous fluids. We then find aerobic germs implanting 
themselves in these fluids, consuming by the aid of 
the oxygen of the air the products of anaerobic fer- 
mentations, and thus completing the mineralization 
of the organic substance. 

Such are the phenomena which supervene in bodies 
left in the air ; they reduce these bodies to pulp in a 
few weeks, more or less rapidly according to the tem- 
perature. 

Putrefaction under water. In water, anaerobes only 
come into play; in running waters the surface of the 
cadaver is constantly being washed and superficial 
microbic invasion thereby prevented. On account of 
the preservation of the elasticity of the skin by con- 
stant bathing the swelling is more intense and more 
uniform. The accumulation of gases causes the body 
to float, and the blood accumulates in the dependent 
parts, in which alone cadaveric patches are produced. 

Putrefaction in the soil. Burial of a cadaver in a 
porous and absorbent soil is followed by absorption 
by the latter of the organic fluids as they are pro- 
duced. From this results a comparative desiccation 
which maintains a certain degree of consistence in the 
body and interferes with microbic pullulation, whilst 
it favors the invasion of fungi. Hence, we see the 
development of molds (penicillium, aspergillus, etc.) 



36 Manual of Veterinary Microbiology. 

which excite a more complete combustion of organic 
matter; on the other hand, the dissemination in the 
soil of the fluid products of putrefaction renders the 
action of the anaerobic germs more general. 

From a practical point of view these facts ought to 
be taken into account, as their rational application 
enables us to limit the intervention of microbes in 
putrefaction, several of which possess pathogenic prop- 
erties and the emanations from which are in all cases 
to be avoided. The earth-system (burial of the cadaver 
in furnace-dried earth) realizes the ideal from this 
point of view. 

Putrefaction is always slower in water and in the 
soil on account of the lower temperature of these 
media. 

3. R6LE IN THE ORGANISM IN THE PATHOLOGICAL CONDI- 
TION. 

A certain number of microbes which meet with 
conditions favorable to their development within the 
organism of animals are the determining cause of dis- 
eases in these animals. These diseases are most fre- 
quently contagious. The pathogenic property of a 
microbe is a functional attribute of its vital faculties ; 
in other words, it is to the life of the germ, to its 
nutritive requirements, to its secretions and excretions, 
and to its multiplication, that we must ascribe the dis- 
turbances which it determines. The study of patho- 
genic microbes, the principal object of this work, will 
be considered in the second and third parts. 



Physiology of Microbes. 



37 



VII. Classification. 

Cohn has proposed to class the bacteria among the 
lower algae. Previously they were generally con- 
sidered as fungi. Several classifications of microbes 
have been proposed, none of which are perfect; we 
reproduce that of M. Guignard. The grouping of 
bacteria, however, has but little importance from our 
point of view. Nevertheless, it will assist the reader 
to a correct understanding of the meaning ascribed to 
certain terms in common use. 



38 



Manual of Veterinary Microbiology. 




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1 


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4 


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K^^> O H^CC 


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.T direction-! 


dividing onl y I 6) filaments, long, divided, without gelatinous matrix . . . . 




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L f isolated or in chains, without gelatinous matri 


a) cells i f in a naked mass 
globular | united 1 in a mass surrounded by a matrix. . 


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the r f d rm f i finanakedmass 

(batonnets) united 1 . 
' L L in a mass surrounded by a matr 
















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f f very short 
RtraiVht J filamentous, without sheat 
\ filamentous, with sheath. 
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Pathogenic Microbes in the Static Condition. 39 



PART SECOND. 

GENERAL CONSIDERATIONS UPON PATHOGENIC 
MICROBES. 

This second part of our work will embrace : 1st. the 
study of pathogenic microbes in the static condition, 
that is to say, considered especially as to their general 
localizations, without taking into consideration their 
action upon the economy ; 2d. the study of the recip- 
rocal reaction of the organism and the microbe, which 
we will designate by the term physiology of pathogenic 
microbes; 3d. the transformations and destruction of 
microbes -in their relations with hygiene and therapeu- 
tics, and 4th. finally, the methods of determination of 
pathogenic microbes. 



CHAPTER I. 



PATHOGENIC MICROBES IN THE STATIC CONDITION. 

1. Conditions of life in external media and within the economy. 
Sources of infection. 2. Distribution of pathogenic germs. 
3. Modes of contagion. Ways of penetration of pathogenic 
microbes. 

Most micro-organisms live in the external world at 
the expense of dead matter ; these have received the 
name of saprogenic or saprophytic germs. 

Some live in either a permanent or adventitious way 



40 Manual of Veterinary Microbiology. 

upon animals and mankind ; they are called pathogenic 
when, by their pullulation, they give rise to diseases. 
These affections have long been known but the study 
of their causes, the infinitely little, is of quite recent' 
date; such diseases are called infectious. Later, we 
will have to distinguish infections properly so called 
from intoxications of microbic origin. 

The term virulence is applied to the collection of 
properties by which pathogenic microbes are able to 
prove detrimental to living beings. This faculty is 
far from being unchangeable ; it may present all de- 
grees of intensity, in accordance with conditions ca- 
pable of modifying the vitality of the germ. 

The term virus is applied to solid, liquid, or gaseous 
vehicles containing pathogenic germs. 

I. Conditions of life of pathogenic germs in external 
media and within the living organism. 

Those germs which cause diseases do not exclusively 
and necessarily live within the economy. Some of 
them can also multiply in dead organic media ; others, 
indeed, only pullulate within the organism incidentally 
the external media being really their natural field of 
multiplication. 

In accordance with these considerations pathogenic 
germs have been divided into three categories : 

a. Contagious obligatory parasites ; 

b. Contagious facultative parasites ; 

c. Non-contagious facultative parasites. 

Contagious obligatory parasites. These are repre- 
sented by microbes which, under natural conditions, 
only reproduce themselves in the living organism and 
are transmitted from one animal to another without 



Pathogenic Microbes of the Static Condition. 41 

undergoing any evolution in the surrounding media. 
Of these, two varieties are distinguished ; 1st. those 
which, little resistant, perish immediately on leaving 
the organism, and the transmission of which must be 
made directly by contact of a diseased subject with a 
healthy subject : syphilis, gonorrhoea, rabies ; 2d. those 
which, more resistant, are preserved for a certain time 
outside of the economy, without however multiplying, 
and may arrive upon healthy subjects by means of vari- 
ous vehicles, contact with a diseased subject not being 
absolutely necessary : measles, variola, scarlatina, diph- 
theria, glanders, tuberculosis. In this case the source 
of the contagious disease resides principally but not 
exclusively in the diseased organism. If the germ is 
met with elsewhere it will be upon objects which have 
been in direct relation with the diseased or its cada- 
ver ; these objects can, moreover, communicate the 
latent germs of which they are bearers to the ordinary 
media : soil, water, or air, in which these germs pre- 
serve their contagiousness for a period of time more 
or less extended. 

Whatever variations there may be in the mode of 
contagion, the contagious obligatory parasite arrives 
upon the healthy organism in the same condition in 
which it left the diseased subject where it had its 
origin. 

Contagious facultative parasites. These live and 
multiply not only within the organism of animals, but 
also outside of the latter, upon dead organic matters, 
in waters, etc.; germs of pyaemia, septicaemias, gan- 
grene, erysipelas, typhoid fever, asiatic cholera. 

The virulence of some of these germs seems even 
to be diminished by their passage through the organ- 



42 Manual of Veterinary Microbiology. 

ism to such a degree that the disease quickly dies out 
if their original virulence is not restored by a return 
to the outer media ; it is thus with cholera, for exam- 
ple, which spontaneously disappears in winter in the 
countries of central Europe, probably because the ex- 
ternal media have become unsuitable to its multipli- 
cation. In the case of cholera we perceive a grada- 
tion toward the miasms ; but whilst in this last the 
microbe exhausts its effects upon the individual which 
harbors it and does not extend beyond it, the bacillus 
of cholera, on the contrary, proliferates during its pass- 
age through the organism of man and thus increases 
the chances of later infection. 

The source of infection, therefore, for germs of this 
kind is twofold : the diseased organism and the in- 
fected media in which the germs pullulate; these two 
sources possess in the same degree the power of be- 
getting the disease, and the opportunities of infection 
will be more frequent than for the germs of the first 
group in as much as the multiplication of the microbes 
in the different media is an important cause of their 
preservation. 

Non-contagious facultative parasites. These live, in 
the normal condition, in the external media, and it is 
only incidentally that they develop within the organ- 
ism of animals. They do not seem in the latter to 
meet with conditions favorable to their vitality, for 
their effects are exhausted completely in the course of 
the disease which they determine, and the disease is 
not transmitted from one subject to another. The 
condition under which it occurs consists always in the 
impregnation of a healthy organism by germs drawn 
directly from external infected media; in short, the 



Pathogenic Microbes in the Static Condition. 43 

disease is bound to the soil : paludic fevers, perhaps 
yellow fever. 

The three classes of germs which we have just been 
considering explain the distinction formerly made be- 
tween the different forms of virus. The first corre- 
sponds to contagions, the second to miasmatic-contagions, 
and the third to the miasms properly so catted. 

Usually we also class with the miasms the toxic 
gases which are disengaged from cesspools, and the 
poison of the expired air. 

Baumgarten has divided pathogenic microbes into 
exogenous and endogenous, according to whether they 
come from the exterior or from the diseased subjects 
themselves. 

II. Distribution of pathogenic germs. 

A certain number of pathogenic germs are ubiquit- 
ous in nature; we meet with them almost every- 
where. Such is the case with the germs of suppura- 
tion and of septicaemias, on account of their excessive 
production and the readiness with which they live in 
the surrounding media. Specific germs are only in- 
cidentally found in such media. 

In the following lines we will review in succession 
the various media which may contain germs noxious 
for the economy, this study being essential to a cor- 
rect understanding of the different methods of con- 
tagion. 

Air. We have seen, in the first part, that the air 
can hold germs in suspension ; it is natural to think 
that among these we may find some which are pos- 
sessed of pathogenic properties. Moreover, it can 
readily be understood that the dust of the streets 



44 Manual of Veterinary Microbiology. 

and accumulations of dirt will yield to the atmos- 
phere, under the influence of numerous agitations 
of the air, particles contaminated with pathogenic 
germs, which may have been brought there in various 
ways. 

It was for a long time believed that the air expired 
by the diseased was a fertile cause of the diffusion of 
pathogenic germs in the atmosphere, but we now 
know that the expired air never contains germs, 
neither have we succeeded in transmitting diseases in 
this way. 

Transmission by the air has been accomplished ex- 
perimentally for a certain number of diseases (sheep- 
pox, charbon, tuberculosis, vaccinia, etc.) by the dis- 
semination of the dried virus of these diseases in the 
atmosphere. 

Under natural conditions the atmospheric germs 
occur in a state of great dilution and hence the 
chances of infection are extremely limited. If we add 
that the air lends itself little or not at all to their 
multiplication, and that they are destroyed more or 
less rapidly by light, oxygen, and dessication, we shall 
see that the danger from the free atmosphere is al- 
most nil. 

The confined atmosphere of inhabited houses con- 
tains more germs than the external air, that of cities 
more than that of the country. 

The confined atmosphere of infected places can be- 
come the carrier of disease germs and can transmit 
certain contagious diseases either by transporting 
these germs into the respiratory passages, or by de- 
positing them upon alimentary matters or on the sur- 
face of wounds. The pyogenic microbes have been 



Pathogenic Microbes in the Static Condition. 45 

met with in surgical wards, the bacillus of Koch in 
those of the tuberculous. These germs come from the 
dried exudates, dressing cloths, etc. 

It is therefore necessary, in places where diseased 
subjects reside, to take precautions against the germs 
of the atmosphere. 

The condition of desiccation of virulent material 
has necessarily a great influence upon the richness of 
the air in pathogenic products, and all the conditions 
which assist in raising the dust are of such a nature 
as to increase the number of these products. Klebs 
has observed that during an epidemic of diphtheria a 
large number of new cases occurred after the sweep- 
ing of the streets and that they especially prevailed 
along the roads followed by the wagons used in trans- 
porting the dirt. 

Waters- From a pathological point of view, gen- 
erally speaking, a water is the more to be feared the 
greater the proportion of organic matter it contains, 
since this material implies the presence of microbes 
which live at its expense. Most of the germs of 
water, however, are inoffensive; only rarely have 
pathogenic microbes been met with : septic vibrio, pus 
cocci, etc. 

Pathogenic germs find entrance to waters in various 
ways ; they may come from the air, from the bodies 
of animals which have succumbed to infectious dis- 
eases, or they may come from the soils traversed by 
the waters. This last mode* has especially attracted 
attention during recent times : the bacillus of typhoid 
fever of man which is voided with the faecal matters, 
can pass with the liquids of cesspools through very 
porous soils and thus come to contaminate the subter- 



46 Manual of Veterinary Microbiology. 

ranean waters. When these waters are used for hu- 
man consumption an epidemic of typhoid fever may 
result. The knowledge of this fact has enabled us in 
numerous cases to trace the disease to its source and 
check its extension. The bacillus of cholera is dis- 
seminated in the same manner. 

Generally speaking, therefore, waters charged with 
organic matters should be viewed with suspicion, and 
it is necessary to take particular precautions in order 
to avoid the pollution of alimentary waters by cess- 
pools, dung-hills, trenches of liquid manure, by the 
stagnant waters of the streets, etc. 

The vicinity of these reservoirs of organic detritus 
to the sources of alimentary waters is always to be 
dreaded, especially when the earth is little adapted to 
filtration ; hence, it is preferable, especially in thickly 
populated centers, to secure water which has filtered 
through virgin soils and convey it to the cities by a 
system of closed canals. 

Pathogenic bacteria preserve their virulence in 
water for varying periods. It has been observed that 
this virulence is preserved during four months for the 
bacillus of charbon, one year for its spores; the bacillus 
of typhoid fever retains its vitality for two months in 
water, that of cholera for twenty-four hours in cess- 
pools, and twenty-nine days in spring water; those of 
glanders and tuberculosis for twenty and ten days re- 
spectively (in water). 

Soil. The germs of the soil are numerous, their 
function being to transform organic matters of which 
this medium is the great recipient ; but most of them 
are without effect on the organism of animals. Never- 
theless, inoculation of vegetable mold into the small 



Pathogenic Microbes in the Static Condition. 47 

animals of the laboratory often rapidly leads to death 
with suppuration or gangrene as local lesions. Wounds 
contaminated by the soil readily become complicated 
by accidents of the same kind. 

The specific germs which are incidentally met with 
in the soil are those of tetanus, charbon, Pasteur's 
septicaemia, typhoid fever, cholera, etc. 

Germs are especially abundant in the superficial 
layers of the soil. The almost constant humidity of 
the soil in winter carries them into the deep layers ; 
on the other hand, during periods of drought, in 
the absence of descending currents of water, the su- 
perficial washing is less complete and the germs re- 
main more in the layers in contact with the atmosphere. 
Thus, other things being equal, the dust will be richer 
in germs in summer than in winter. 

The micro-organisms of the soil become harmful in 
various ways : 1st. by contaminating the vegetable 
foods which grow . upon an infected place ; 2d. by 
distributing themselves in the air through the desicca- 
tion of the soil ; 3d. by directly contaminating a so- 
lution of continuity (tetanus) ; -4th. by contaminating 
the waters which filter through the earth and which, 
later, are to serve for alimentation. 

In the first three cases the germs must occupy the 
surface of the arable layer ; microbes which are more 
deeply situated may return to the surface by the in- 
crease in height of the sheet of subterranean water, 
by the phenomenon of capillarity which constantly 
occurs in finely divided soils, or, finally, through the 
intermediation of earth worms, larvae, etc., a fact 
which has been demonstrated by M. Pasteur in the 
case of charbon. 



48 Manual of Veterinary Microbiology. 

The infection of subterranean waters by the soil is 
subject to numerous influences dependent upon the 
location and nature of the land. Changes of eleva- 
tion of the surface waters also produce effects which 
it is of importance to consider. It has been noticed 
that large floods are often followed by typhoid epi- 
demics, a circumstance which has been attributed to the 
simultaneous rise in level of the waters of infiltration 
generally, and especially adjoining large sheets of 
surface water ; this elevation of the subterranean wa- 
ters brings them into contact with soils impregnated 
with putrid matters in the neighborhood of cesspools, 
trenches of liquid manure, sewers, etc., and when 
these putrid matters contain the typhoid germ there 
results a general pollution of springs and wells into 
which these contaminated waters diffuse themselves. 

The pathogenic germs of the soil can retain their 
vitality for a period more or less extended ; Grancher 
has seen the typhoid bacillus retain its vitality in the 
soil for five months. The charbon bacillus is also pre- 
served in this medium, more especially, however, in 
the form of spores. 

The destruction of these germs is dependent upon 
the action of the oxygen and light ; as the conditions 
of their pullulation are more delicate than those of 
saprogenic microbes, the presence of the latter must 
also be taken into consideration in so far as their ac- 
tive proliferation more or less rapidly brings about an 
insufficient supply of nutrition for the pathogenic 
species. 

Foods. The vegetable foods may be contaminated 
by pathogenic germs either by contact with the soil 
while yet in growth (charbon, actinomyces), or, after 



Pathogenic Microbes in the Static Condition. 49 

harvesting, in the storehouses of fodders by the air 
of infected stables or barns, or from direct contact 
with sick animals. 

MM. Galtier and Violet attribute to the fodder the 
development of the typhoid affections of the horse. 

Foods of animal origin are sometimes bearers of 
pathogenic germs. This is the case when they come 
from subjects suffering from diseases caused by these 
germs. The flesh and all the tissues of animals dead 
from charbon contain the bacteridium. 

Dwellings and vehicles. Places which have been oc- 
cupied by animals affected with contagious diseases, and 
vehicles (wagons, etc.) which have been used in their 
transportation are most frequently infected by specific 
germs. These are deposited upon the floor, walls 
and ceilings of houses and upon the mangers, racks, 
etc., either by the air or directly with the secretions or 
excretions of the sick. The transmission of microbic 
diseases through the intermediation of dwellings is 
therefore in every way possible. 

Various articles and utensils. All the articles and 
utensils which are found in places occupied by diseased 
animals or which have been employed in the service 
of the latter harness, blankets, grooming instru- 
ments, sponges, brushes, curry-combs, etc., litter and 
manure may become bearers of pathogenic germs 
which have been derived from these animals. 

Healthy organism. The bodies of animals in health 
are always bearers of microbes ; but the latter are 
most frequently of no importance from our present 
point of view. We already know that the ubiquitous 
pathogenic germs those of suppuration and of septic 
5 



50 Manual of Veterinary Microbiology. 

accidents which are met with almost every- where 
must be present on the surface of the body and even 
in its interior, upon the various mucous membranes 
and especially that of the digestive canal. Recently 
*the bacillus of tetanus has been discovered in the 
fsecal matters of healthy horses. The pneumococ- 
cus the pathogenic germ of lobar pnuemonia in 
man is constantly present in the buccal mucus of 
nearly all individuals. The skin, the respiratory mu- 
cous membrane and that of the genito-urinary pas- 
sages near the openings communicating with the ex- 
terior, are likewise contaminated with ubiquitous 
germs. 

The transmission of a contagious disease through 
the intermediation of healthy individuals, animals or 
mankind, shows that pathogenic micro-organisms 
may be present on or in the body without producing 
disease. The methods by which infection is produced 
and the phenomenon of immunity sufficiently explain 
this peculiarity. 

Infected organism. The organism attacked by a 
contagious disease harbors the microbes of the latter 
in very different points according to the nature and 
localization of that disease. It is desirable, for the 
ends of a rational prophylaxy, to know the places of 
election of the germs of the different diseases, and, 
more especially, the seat of those which, in being 
eliminated from the economy, are able to contaminate 
healthy individuals. 

We may meet with these germs in the various 
secretions flowing to the exterior: in the saliva: 
rabies; in the faecal matters: tuberculosis, typhoid 
fever, cholera, chicken cholera, pneumo-enteritis ; in 



Pathogenic Microbes in the Static Condition. 51 

the expectorations : tuberculosis, glanders, actinomy- 
cosis, sheep pox; in the urine: bacterial hsemoglobinu- 
ria of cattle, tuberculosis, etc.; in the semen and vagi- 
nal mucus : gonorrhoea ; in the secretions of wounds : 
glanders-farcy ulcers, syphilitic chancres, lesions of 
douriue, pustules of variola of the different species of . 
animals. 

Hemorrhages may occur through the various pas- 
sages and thus distribute externally the germs which 
are present in the blood : charbon. 

If germs do not appear to be eliminated by the in- 
tact skin it is none the less true that the latter is fre- 
quently soiled by pathogenic germs emitted by the 
diseased; these germs are conveyed to the skin 
through contact of the secretions or infected litters. 

A subject affected with a contagious disease does 
not appear to be at all stages of the disease capable 
of communicating the contagion to the same degree. 
Thus, in the case of glanders and tuberculosis, the 
danger appears to be absent if the softened lesions do 
not directly communicate with the exterior. 

Cadavers of infected subjects. The bodies of animals 
dea*d of infectious diseases are fertile sources of patho- 
genic germs; from the stand-point of the alimentary 
hygiene of man it is of great importance to recognize 
the place of election of those germs, but the question 
can hardly be considered in a general manner. 

As to the duration of the virulence of pathogenic 
microbes in dead bodies left to themselves, it varies 
greatly according to the germs concerned, but upon 
this subject our knowledge is very incomplete. We 
know, however, that in some diseases the pathogenic 
power may persist for a long time for years in the 



52 Manual of Veterinary Microbiology. 

case of charbon and tuberculosis, for example. It is 
therefore necessary to carefully destroy these bodies. 

III. Modes of contagion. Ways of penetration of patho- 
genic microbes. 

Those germs which are exciters of disease, after 
having multiplied in the bodies of the first infected 
animals, may be transported to other animals, and 
thus propagate the disease. This transference of the 
germs of a disease from a sick to a healthy individual 
constitutes contagion. 

1. Modes of contagion. A microbic disease will the 
more surely be communicated, the contagion will have 
greater opportunity of taking effect, in proportion to 
the number of the germs emitted by the diseased. A 
superficially situated disease will be more readily trans- 
missible, other things being equal, than a deep seated 
one ; a disease with lesions of large extent will be 
more easy to communicate than one in which these 
are less extensive. The greater or less duration of the 
resistance of the virus to the natural agents of destruc- 
tion is also a condition upon which depends the aug- 
mentation or diminution of the contagion-begetting 
power of the virus. 

The transference of pathogenic germs from a 
diseased subject to a healthy subject contagion may 
be direct or immediate, that is, the healthy subject ob- 
tains the germs of the disease from the diseased ani- 
mal itself, or it may be indirect or mediate, the healthy 
individual receiving the microbes eliminated by the 
diseased, through the intervention of external media. 

Direct or immediate contagion. Of this we have to 
distinguish two cases according as the transmission 



Pathogenic Microbes in the Static Condition. 53 

takes place between two subjects independent of each 
other (direct contact), or from the mother to the foetus 
(heredity). 

Direct contact. Direct contact seems to be necessary 
for a certain number of diseases: rabies (bites), syph- 
ilis, dourine (coition), in which the germs, conta- 
gious obligatory parasities, appear not to maintain 
their virulence in the surrounding media. On the 
contrary, in other diseases direct contact is not essen- 
tial, the microbes maintaining their virulence outside 
of the organism : glanders, tuberculosis, charbon, etc. 

Man becomes infected by direct contact when he 
accidentally inoculates himself in making autopsy on 
bodies affected with glanders, tuberculosis or charbon. 

Heredity. Contagious diseases appear to be trans- 
mitted from the mother to the foetus only by passage 
of the specific microbes through the placenta. The 
intervention of the father in phenomena of this kind 
is therefore indirect, in as much as the disease with 
which he is affected must first be transmitted to the 
mother, a fact which is observed in syphilis, for ex- 
ample. Here, therefore, the action of the mother in 
reality alone comes into play. 

Some infectious diseases are transmitted from the 
mother to the foetus; these are especially general af- 
fections and those in which the germs are able to .cir- 
culate in the blood : certain septicaemias, charbon, 
fowl-cholera, strangles, rouget, tuberculosis. 

The transmission seems to occur through alterations 
of the placenta, such as hemorrhages, specific lesions 
(tubercles, etc.), alterations which, indeed, are readily 
produced on account of the diseased condition of the 



54 Manual of Veterinary Microbiology. 

mother and the tendency of the various germs living 
in the blood to determine vascular and other lesions. . 

Nevertheless, this interpretation, which certain ob- 
servations sufficiently justify, does not explain all the 
facts of heredity, and it seems that a simpler mechanism 
intervenes in some cases. 

We know that the contagious affections of the geni- 
tal apparatus of the female are readily transmissible, 
and they will actually be transmitted to the foetus un- 
less their existence renders gestation impossible, or 
provokes abortion. 

The disease transmitted to the foetus may cause the 
death of the latter and its premature expulsion, it may 
disappear, or, finally, may remain in a stationary con- 
dition, permitting the complete development of the 
young animal, in which, at a later period, it may un- 
dergo fresh evolution. 

Hereditary transmission may be limited to the com- 
munication of immunity by the diffusion through the 
placenta of soluble vaccine substances elaborated 
within the body of the mother. But immunity in the 
young being may also be consecutive to recovery from 
the disease with which it has itself been affected. 

Indirect or mediate contagion. We call the contagion 
indirect when the virulent germs which come from 
diseased subjects are transported on to healthy indi- 
viduals after having been deposited on its surround- 
ings. We have already seen in considering the dis- 
tribution of pathogenic germs in external media, that 
they may be encountered in the air, water, the soil, 
foods, the walls of houses, on mangers, racks, manure, 
litter, and the various objects and utensils which have 
been more or less directly in contact with the diseased. 



Pathogenic Microbes in the Static Condition. 55 

!N"ow, these are just the infected vehicles which trans- 
port the pathogenic germs to healthy individuals. 

Indirect contagion only occurs with facultative 
parasitic germs and with contagious obligatory para- 
sitic germs which are endowed with considerable 
power of resistance against external causes of de- 
struction. 

The point of entry of microbes mediately trans- 
mitted is variable; when the virulent matters are 
brought into contact with the skin, or with the 
genito-urinary or ocular mucous membranes, it is 
called transmission by indirect contact; when the con- 
tagion is introduced with the food, water, or by 
the air, it is called transmission by ingestion or inhala- 
tion. 

When pathogenic germs are once deposited on or 
within the organism, the manner in which they may 
aifect the latter will differ according to the case; 
sometimes they will have no appreciable effect; at 
other times they may determine the irruption of a 
disease similar to that which has engendered them. 
The placing in activity of the pathogenic faculty de- 
pends in reality upon many circumstances, and, in 
the first place, upon absorption. We will study here 
the ways and processes of absorption of pathogenic 
microbes, as a sequel to the study of modes of con- 
tagion. 

2. Absorption of pathogenic microbes. Disease germs 
are capable of penetration through various surfaces, 
natural or artificial; these we will now review in suc- 
cession. 

Skin. The intact skin is an unfavorable surface 
for the absorption of germs, but does not oppose it- 



56 Manual of Veterinary Microbiology. 

self in an absolute manner to their penetration; some 
of them are probably able to introduce themselves 
into its substance through the pilo-sebaceous glands, 
and thus give rise to diseases. The anatomical pus- 
tules which develop upon the hand or arm of anato- 
mists, surgeons, and accoucheurs, seem to be pro- 
duced in this way ; it is the same with acne. 

Repeated frictions of the skin at the time of con- 
tact with virulent matters will considerably further 
penetration. Garre induced the formation of furun- 
culous pustules on his own arm by rubbing it with a 
culture of staphylococcus pyogeues aureus. 

The incorporation of the virus with a fatty body, 
by rendering the contact more complete, increases 
the facility and certainty of absorption by the intact 
skin. Charbon and glanders have been communi- 
cated in this way. 

In short, if absorption by the healthy skin is pos- 
sible, it seems to take place only within narrow limits. 
It acquires, on the other hand, great importance 
when a traumatism has opened the way to microbes, 
most of these being capable of penetrating through 
even the slightest solutions of continuity in the ex- 
ternal integument. 

Digestive mucous membrane. The different structure 
of this membrane in the various parts of the digest- 
ive canal manifestly implies variation in its absorp- 
tive faculty toward pathogenic microbes. The lining 
membrane of the anterior passages is little adapted 
to their penetration, but this much more readily oc- 
curs if one or several accidental solutions of con- 
tinuity exist in these passages. Experiment has 
shown that the addition to fodders contaminated 



Pathogenic Microbes in the Static Condition. 57 

with the virus of charhon, of bodies capable of ex- 
coriating the mucosa (thistles, husks of barley) in- 
creases the mortality from this disease, but excoria- 
tions of the bucco-pharyngeal mucosa are compara- 
tively common, and consequently it is necessary to 
take into account this way of infection. 

Absorption, however, appears occasionally to take 
place in this situation in spite of the integrity of the 
mucosa, as, for example, when we contaminate a 
healthy flock with aphthous stomatitis by depositing 
a little of the saliva coming from diseased animals in 
the mouth of other healthy animals. 

As in the case of the anterior, the mucous mem- 
brane of the posterior digestive canal admits of pen- 
etration by virus when it is the seat of alterations in 
its continuity : erosions, ulcerations. But, even when 
it is intact, the gastro-intestinal mucous membrane 
does not prevent the absorption of pathogenic germs, 
as numerous experiments have demonstrated. Char- 
bon, glanders, tuberculosis, aphthous fever, cholera 
of fowls, and, indeed, nearly all microbic diseases can 
be transmitted in this way. Nevertheless, all sub- 
jects which ingest virulent products do not neces- 
sarily become infected ; a certain number are refrac- 
tory to the disease; in others the gastric juice kills 
all the germs which are introduced, whilst in others, 
again, absorption may not occur. 

Referring to the microbicidal action of the gastric 
juice, we may repeat, that non-sporulated bacteria 
are killed much more rapidly than spores ; infection 
by the latter is consequently infinitely more certain ; 
this fact has been demonstrated for charbon. 

Respiratory mucous membrane. Air charged with 



58 Manual of Veterinary Microbiology. 

virulent dust, being inhaled by a healthy subject, may 
determine the outbreak of the disease, this fact hav- 
ing been established for tuberculosis, charbon, gland- 
ers, fowl cholera, etc. The experimental injection of 
microbes within the trachea gives the same result. 

The respiratory mucous membrane, therefore, allows 
itself to be traversed by microbes. Nevertheless, the 
presence of pathogenic germs in the air does not im- 
ply that infection will necessarily take place. These 
germs are generally deposited upon the mucus of the 
anterior passages and are then rejected with the prod- 
ucts of expectoration. The fact that germs are con- 
stantly absent from the expired air whilst they are 
always contained in the inspired air, sufficiently shows 
that the latter is purified in contact with the mucous 
membrane. 

Absorption by the respiratory mucous membrane 
may occur throughout its whole extent; as with other 
lining membranes, this absorption is favored by solu- 
tions of continuity. 

Ocular mucous membrane. The conjunctival mu- 
cous membrane absorbs certain microbes; of this, 
accidental inoculation of gonorrhoeal pus in the eye 
furnishes sufficient evidence. M. Galtier has suc- 
ceeded in transmitting rabies in this manner. 

Genito-urinary mucous membrane. Syphilis, gonor- 
rhoea, in man, and dourine in the horse are generally 
inoculated by coition in the absence of solutions of 
continuity either of the vagina or urethra. Fric- 
tions probably aid in the penetration of the specific 
germs. Absorption by these intact passages can, 
therefore, not be doubted. 

The penetration of microbes, therefore, takes place 



Pathogenic Microbes in the Static Condition. 59 

through the intact mucous membranes, but this faculty 
may give rise to no injurious effects upon individuals 
who are in absolutely physiological conditions. M. 
Bouchard has given the following explanation of the 
mechanism by which the normal organism opposes 
itself -to the penetration of the germs present on the 
natural surfaces: Germs which have traversed the 
epithelium immediately come into conflict with the 
white corpuscles distributed in the derm of the mu- 
cosa, and by these they are taken up and digested. 
If, on the contrary, the economy is disturbed, thrown 
out of equilibrium, by a sufficient cause (repeated in- 
fluence of currents of cold air, influence of fear) the 
enfeebled white corpuscles allow the microbes to 
break through the barrier which they are charged 
with defending. The experiments of Bouchard were 
made with ordinary germs ; their results are applica- 
ble to pathogenic microbes, with this difference, that 
it is necessary, here, to take into account a supple- 
mentary factor directly related to the pathogenic 
faculty : we refer to the influence that the secretions 
of these microbes can themselves exert, in such cases, 
upon the white corpuscles, in attenuating or annihi- 
lating their action. 

Wounds. In general, wounds are the surfaces of 
predilection for the absorption of pathogenic germs 
not only on account of the division of the tissues and 
vessels which, in a manner, opens the way for them, 
but also because the organism is not prepared with 
its means of defense. The latter has to be organized 
upon the field, and but too frequently proves inade- 
quate to repel the invaders. Nevertheless, hemor- 
rhage, and the phagocytic action of the elements of 



60 Manual of Veterinary Microbiology. 

the tissues, as well as that of the leucocytes which 
speedily accumulate in the wound, are conditions un- 
favorable to absorption. The latter, moreover, de- 
pends upon many other circumstances bearing upon 
the nature of the germ, its vehicle, the depth and 
extent of the wound, etc. The microbes of tetanus 
and those of gaseous gangrene, for example, only 
multiply in wounds to which the access of the air is 
limited; their activity is checked by atmospheric 
oxygen. The rapidity of penetration is influenced 
by the nature of the medium ; an aqueous medium 
will be more readily absorbed than a solid excipient 
or one of thick, colloid consistence. 

Tuberculosis, symptomatic charbon, and grangren- 
ous septicaemia are not inoculable by sub-epidermic 
punctures, whilst this inoculation is successfully per- 
formed in the subcutaneous cellular tissue. Absorp- 
tion is always more easy when the tissue is itself 
lacerated. 

Absorption from wounds is, in general, very rapid ; 
glanders has been seen to supervene in spite of deep 
cauterization of the inoculated wound two hours after 
the insertion of the virus ; cauterization after a lapse 
often minutes has still allowed the evolution of sheep- 
pox. The amputation of the ear of a rabbit inocu- 
lated with charbon by sub-epidermic puncture in that 
region has not prevented the irruption of the disease, 
although this operation followed only three minutes 
after the inoculation. 

Infection by wounds may be local only, or it may 
become generalized; in the latter case the extension 
occurs chiefly by the lymphatics, the germs then 
showing their presence in these vessels by the lesions 



Physiology of Pathogenic Microbes. 61 

which they determine in the corresponding glands. 
But pathogenic germs can also penetrate directly 
into the blood-vessels in which case generalization 
occurs much more rapidly. 



CHAPTER II. 

PHYSIOLOGY OF PATHOGENIC MICROBES. 

1. Action of pathogenic microbes upon the organism. Receptiv- 
ity. Immunity. 2. Reaction of the organism against path- 
ogenic microbes. Phagocytosis. Bactericidal condition. 
3. Evolution of the bacterial disease. 

I. Action of microbes upon the organism. 

MECHANISM OF THE PATHOGENIC ACTION. 

Pathogenic germs exert their action upon the 
economy in two principal ways, of themselves, or by 
their secretion products. We will consider, suc- 
cessively, the mode of development of the troubles 
which they occasion, both local and general. 

1. Pathogeny of local alterations. In local lesions 
the microbe acts at first like a foreign body, that is, 
it excites a purely mechanical irritation; the Koch 
bacillus very' probably acts in this way when it gives 
rise to tubercular neoplasms ; we know, indeed, that 
the injection of lycopodium powder into the blood 
develops a similar lesion, and the egg of the strongy- 
lus vasorum appears to act in the same way in the 
pseudo-tubercles which it occasions in the dog. 



62 Manual of Veterinary Microbiology. 

This mechanical influence, however, seems to be 
much the least important. The soluble substances- 
diastases and ptomaines secreted by the microbes 
possess very active properties and to these we ascribe 
the genesis of most of the manifestations of the in- 
fectious diseases. 

Microbic diastases, as we have already said, repre- 
sent the digestive juices of the microbes; they can 
therefore act chemically upon the substance of the 
tissues, provoking hydrations and chemical decom- 
positions, and thus lead to the liquefaction, softening, 
and even mortification of these tissues. These dis- 
solutions are sometimes preceded by a sort of coagu- 
lation (coagulation necrosis). 

The soluble substances diastases and ptomaines 
may possess phlogogenic properties; in this case 
they develop, of themselves alone, all the inflamma- 
tory phenomena which follow the inoculation of the 
microbes from which they come. They excite swell- 
ing, proliferation, and fatty, hyaline, or waxy de- 
generation of the elements, dilatation of the vessels, 
exudation of products more or less plastic, sometimes 
active diapedesis of the white blood corpuscles and 
even extravasations. The pneumobacillus liquefaciens 
bovis, isolated by Arloing from the lesions of bovine 
contagious pleuro-pneumonia, secretes a diastase 
which excites inflammatory oademas recalling those 
of the disease itself. The staphylococcus pyogenes 
aureus produces a diastase and a ptomaine which are 
phlogogenic and pyogenic. 

The local alterations which supervene in conse- 
quence of the introduction of certain germs into the 
tissues sometimes depend, at least in part, on the 



Physiology of Pathogenic Microbes. 63 

chemical action to which the nutrition of these 
germs gives rise; the emphysernatous tumors of 
symptomatic charbon and of traumatic gangrene are 
caused by the abnormal production of gas which 
accompanies the fermentations provoked by the 
anaerobic microbes of these diseases. 

2. Pathogeny of remote and general manifestations. 
Anatomical alterations remote from the original point 
of infection may be produced as a consequence of the 
penetration of the pathogenic microbes into the lym- 
phatics and blood-vessels ; their pathogeny is identi- 
cal with that of the local manifestations. 

The arrest of the microbes in vessels of small cali- 
ber may become the starting point of secondary me- 
chanical lesions; we refer to microbic embolisms, 
comparatively frequent in general infectious diseases, 
and which are followed by infarcts, stases, etc. But 
it may happen that the secretion-products alone pene- 
trate into the circulation, the bacteria remaining in- 
trenched at the primary focus; we may also have 
localized alterations dependent upon special proper- 
ties, dissolving, phlogogeriic, or pyogenic, of these 
products. 

Besides the functional troubles resulting directly 
from the anatomical lesions, primary or secondary, 
seated in the various organs, we have to consider 
the genesis of the general manifestations which ac- 
company microbic diseases, either local or general. 

Fever is one of the most frequent symptoms ; a 
certain number of soluble microbic substances excite 
hyperthermia. This almost always results from a gen- 
eral nutritive excitation of the tissue elements, from 
contact with these substances ; but it may also be the 



64 Manual of Veterinary Microbiology. 

result of a diminution in the loss of heat through 
constriction of the peripheral capillaries. Rise of 
temperature therefore does not, in all cases, imply 
increased production of heat. 

Several microbic substances are known to possess 
this pyretogenic or fever-begetting property : those 
derived from the bacillus of blue pus, from Freid- 
lander's microbe, etc. 

Further, the soluble products of the anatomical 
elements themselves produce similar effects. Ac- 
cording to Gangolphe and Courmont, necrobiosis of 
the tissues develops pyretogenic substances inde- 
pendent of all microbic intervention. These authors 
have observed that bistournage is followed by fever 
when the testicular products are able to penetrate 
into the blood, whilst fever is absent if the precaution 
be taken to put a ligature completely around the scro- 
tum so as to prevent all absorption. It is not impos- 
sible that the -super-activity of the phagocytes, strug- 
gling against microbic invasion, becomes also the 
starting point of the production of pyretogenetic 
substances, a hypothesis which certain facts seem to 
indicate. "We know, moreover, that extracts of flesh 
and of the spleen possess similar properties, which 
thus appear to belong to a certain number of the 
residues of normal disassimilation. 

Some soluble substances of microbic origin excite 
the phenomenon of hypothermia, lowering the tem- 
perature of the body ; this property is possessed by 
the soluble products of the comma bacillus of cholera, 
the septic vibrio, and the staphylococcus aureus. 

The nervous system is especially sensitive to the 
action of these products of microbic origin, in the 



Physiology of Pathogenic Microbes. 65 

way of excitation (hyperkinesia developed by the 
toxines of tetanus), or of depression (coma, somno- 
lence, in fowl cholera and charbon ; paralysis consecu- 
tive to diphtheria and to the pyocyanic disease). The 
heart, the respiratory centers, and the vaso-motor cen- 
ter may also feel the influence of these substances, 
from which may result sometimes an increase, some- 
times a diminution, of the functional activity of these 
organs. 

RECEPTIVITY. 

Receptivity, or aptitude to contract infectious dis- 
eases, varies in accordance with a large number of 
circumstances, and, especially, with- the species and 
the mode of inoculation, with the individual, age, 
heredity, causes of depression, the quantity and the 
quality of the virus, and the association of the virus 
of more than one disease. 

1. Influence of species and of the mode of inocula- 
tion. The susceptibility of a given animal species 
to a disease, when experimentally inoculated, does 
not necessarily imply the liability of that species to 
contract the disease spontaneously. 

Tuberculosis develops spontaneously, with great ease 
and frequency, in man, cattle, and birds ; it is much 
rarer in the horse, the pig, and the dog under the 
same conditions; as for the rabbit and the guinea 
pig, which it is very easy to inoculate experimentally, 
they do not contract the disease except by inocula- 
tion. 

Symptomatic charbon, appearing spontaneously 
only in bo vines, is inoculable to the sheep, the goat, 
and the guinea pig, but not to other animals. 
6 



66 Manual of Veterinary Microbiology. 

Rabies, a spontaneous* disease of the dog, is inoc- 
tilable to all species of mammals and to birds, intra- 
cranial inoculation being always successful, whilst 
hypodermic inoculation gives .only variable results. 

Bacteridian charbon develops naturally in cattle, 
sheep, and horses, experimentally in all the domesti- 
cated mammals. 

Influence of the individual. All the individuals of 
any species susceptible of contracting a microbic 
disease do not take that disease when they are ex- 
posed to the contagion; some of them are less sus- 
ceptible than others and a few may even be absolutely 
refractory. 

Influence of age. Strangles, and distemper of dogs 
are diseases of youth; symptomatic charbon only ap- 
pears in cattle of from six months to four years and 
is with difficulty inoculated to the young calf ; guinea 
pigs are the more susceptible to charbon the younger 
they are. Pigs of less than four months are much 
less susceptible to rouget than adults; hence, only 
very young pigs should be vaccinated against this 
disease. 

4. Influence of heredity. Certain predispositions are 
inherited ; for example, that of children born of con- 
sumptive parents, to contract tuberculosis. 

5. Influence of depressive causes. All conditions 
which have a debilitating effect on the organism fa- 
cilitate microbic invasion. Pasteur has shown that 
lowering the temperature of fowls inoculated with 
charbon is followed by the evolution of the disease in 
these animals in spite of their natural immunity. 

* [Acquired by transmission from its own kind. D.] 



Physiology of Pathogenic Microbes. 67 

Bouchard has shown that gradual cooling of the 
guinea pig allows the entry into the blood of this 
animal of the microbes distributed on the surface of 
the mucosa. Nervous disturbances of a depressive 
character act in the same way : fear. 

The administration of chloral, curara, or alcohol to 
the frog and to the dog, which are refractory to bacte- 
ridian charbon, endows them with receptivity for this 
disease. Antipyrine and chloral diminish the resist- 
ance of the chicken to the same affection. 

The attenuated and inoffensive bacillus of symp- 
tomatic charbon becomes pathogenic and causes the 
disease when it is conjoined with lactic acid, or tri- 
methylamine, or when injected into a tissue pre- 
viously contused. The same substances, as well as a 
similar traumatism, also favor the implantation of 
the tetanus bacillus. 

The influence of these very diverse conditions 
must be ascribed to the action which they exert upon 
the phagocytes, the protective function of which they 
diminish, by rendering them less capable of digesting 
the microbic invaders. 

6. Influence of the quality and quantity of the virus. 
Receptivity naturally varies with the special degree 
of virulence of the germs that the organism receives; 
a germ which is toxic for a given species may become 
an efficacious vaccine when its virulence has been en- 
feebled. 

As to the quantity of the virulent substance, it 
especially requires attention ; the influence of the 
dose can not be questioned ; the natural immunity of 
Algerian sheep against charbon is overcome by inoc- 
ulation of a large dose of the charbon bacillus. In- 



68 Manual of Veterinary Microbiology. 

oculation of a minimum quantity of certain germs 
confers immunity, whilst a larger dose produces the 
fatal disease (gangrenous septicaemia, symptomatic 
charbon). 

7. Influence of microbic associations. The suscepti- 
bility to an experimental disease is increased by in- 
jecting, at the beginning of this disease, a consider- 
able amount of the soluble products coming from the 
microbe inoculated (Bouchard). This fact has been 
established for the charbon bacteridium, fowl cholera, 
the staphyloccus aureus, bacillus prodigiosus, the ba- 
cillus of symptomatic charbon, the pyocyanic bacil- 
lus, etc. This is the more striking since the injection 
of the same products without microbes often confers 
immunity. Here, on the other hand, it aggravates 
the trouble or renders it possible, by overcoming the 
natural or acquired immunity. The mode of action 
of these substances consists, in this case, in the ob- 
struction which they oppose, by paralyzing the vaso- 
dilator nerves, to the diapedesis and phagocytosis 
which the germs of the disease naturally excite when 
they are inoculated to vaccinated subjects, or those 
which are naturally refractory. 

The adjuvant action that the microbic secretions 
exert in association with the microbe from which 
they come, may also be manifested in association with 
other germs ; thus, the natural immunity of the rab- 
bit against symptomatic charbon is obliterated if we 
inject to this animal, at the same time with the char- 
bon bacteria, a sterilized culture of the staphylococcus 
aureus, or micrococcus prodigiosus. 

The same interpretation is applicable to the predis- 
posing or aggravating influence that a previous or 



Physiology of Pathogenic Microbes. 69 

concomitant disease exerts upon the course of an- 
other infectious disease. 

In the case of tetanus, however, the association of 
accessory germs, such as the bacillus prodigiosus, 
renders more certain the irruption of the disease, by 
provoking diapedesis, because the medium then be- 
comes more favorable to the multiplication and nu- 
trition of the tetanus bacillus. 

On the other hand, the presence of phlogogenic 
germs, which excite an excessive diapedesis, may op- 
pose the receptivity for a given microbe. Thus, re- 
covery from malignant pustule nearly always occurs 
after the appearance of suppuration ; this is explained 
by the fact that the white corpuscles, whose accumu- 
lation is brought about by the foreign germs, over- 
come the bacteridium. 

Eeceptivity of the tissues and organs. Pathogenic 
germs do not act in the same manner upon all parts 
of a susceptible organism ; they appear to have a sort 
of affinity for certain organs, for certain tissues. 
Thus, the bacillus of bacteridian charbon multiplies 
in the blood whilst that of symptomatic charbon only 
develops in the connective and muscular tissue and 
is, on the contrary, killed in the blood. 

The pneumococcus generally limits its field of 
operations to the lung; the bacillus of pneumo- 
enteritis to the lung and the intestine; that of glan- 
ders to the respiratory apparatus and to the skin, etc. 

This elective action of pathogenic microbes has, in 
some cases only, received a scientific explanation. 

When the germs have penetrated into the blood 
they will localize themselves preferably in the organs 



70 Manual of Veterinary Microbiology. 

predisposed, enfeebled, hence less effectively prepared 

for defense. 

IMMUNITY. 

Immunity consists in the absence of receptivity; 
it may be natural or acquired. Acquired immunity is 
most frequently consecutive to a first attack of the 
disease. A large number of microbic affections do 
not recur; they leave behind them organic changes, 
inappreciable directly, but more or less permanent, 
which oppose themselves to the later development of 
their germs ; the individual, once recovered, is vacci- 
nated. This is the case in variola, vaccinia, sheep 
pox, syphilis, charbon, rouget, fowl cholera, pleuro- 
pneumonia, etc. Other diseases, on the contrary, can 
reappear several times in the same individual ; such 
are gonorrhoaa, simple chancre, diphtheria, erysipelas, 
tuberculosis, glanders, etc. In the case of these two 
last diseases we see, in reality, specific lesions super- 
vene at periods very remote from the time of the first 
appearance of the affection, which would not happen 
if one or more first attacks of the disease had vacci- 
nated the organism. Immunity may also be com- 
municated by artificial means, and the process by 
which this is effected is known as vaccination. 

The duration of acquired immunity varies greatly 
according to the disease and the individual concerned. 
In general, immunity consecutive to the natural dis- 
ease is more permanent than that conferred by vacci- 
nation. 

Patient researches have thrown a little light upon 
the nature of immunity, and shown that it operates, 
at least in part, through the active intervention of 
the organism which possesses it. 



Physiology of Pathogenic Microbes. 71 

It was at first supposed that the non-recurrence of 
infectious diseases was due to the fact that the mi- 
crobes, at the time of the first attack, had abstracted 
from the blood principles indispensable to their 
growth, or, on the other hand, contaminated the 
blood with principles which opposed their growth. 
In short, two theories were entertained : that of ex- 
haustion and that of impregnation. 

The first is to-day abandoned ; immunity has, in 
fact, been overcome by employing large doses of 
virus, which would be impossible if the organism 
was .really impoverished in substances indispensable 
to the microbes. 

The doctrine of impregnation, on the other hand, is 
strongly supported by the discovery of vaccinating 
substances. A certain number of microbes secrete sub- 
stances which mix with the fluids of the tissues, dif- 
fuse through all the economy, and impair the vitality 
even of the germs which have produced them ; these 
preventive substances which appear not to be identical 
with the toxic secretions of microbes oppose them- 
selves to a recurrence of the disease. 

The existence of soluble vaccinating substances has 
been unquestionably established for a certain number 
of diseases : the blue pus disease, bacteridian charbon, 
symptomatic charbon, cholera, Pasteur's septicaemia, 
pneumo-enteritis of the pig, etc. 

Impregnation of the blood does not account for the 
persistence of immunity. Substances prejudicial to the 
growth of the germs must gradually become elimi- 
nated from the economy, and if vaccination were due 
solely to their presence, its effects would be extin- 
guished in a comparatively short time. 



72 Manual of Veterinary Microbiology. 

The doctrine of the impregnation of the humors is 
happily supplemented by that of the modification 
of the solid parts of the organism, the anatomical 
elements. 

In acquired immunity, whether it be consecutive to 
a first attack of the disease or be conferred by vac- 
cination, the fluids and tissues possess bactericidal 
properties. Thus, the serum of the rabbit vaccinated 
against the pyocyanic disease, is bactericidal for the 
microbe of this disease; but this property also be- 
longs to the solid tissues of the vaccinated animals, a 
fact which has been further demonstrated by the fail- 
ure of attempts to cultivate the bacillus of symptom- 
atic charbon in the thigh of a guinea pig vaccinated 
against this affection. The production of the bacteri- 
cidal state by vaccination is established for a certain 
number of germs : the bacillus of bacteridian char- 
bon, symptomatic charbon, blue pus, the vibrio of 
cholera, and MetschnikofFs vibrio. 

This property is communicated to the vaccinated 
animal by the mingling of the vaccinating substances 
with the nutritive liquids of the economy ; the con- 
tact of these fluids with the tissue elements brings 
about a permanent nutritive modification of the lat- 
ter; this modification, throughout all the time of the 
duration of the immunity, exerts its influence upon the 
fluids of the body, endowing them with the microbi- 
cidal faculty. By virtue of this faculty the virulent 
germs against which the vaccination has been directed, 
when they attempt to invade the organism, find them- 
selves opposed, attenuated, through contact with the 
fluids, their secretion products which tend to diminish 
diapedesis are less abundantly produced and the phag- 



Physiology of Pathogenic Microbes. 73 

ocytes, as a result of the reaction which naturally 
occurs on contact with foreign bodies, issue from the 
vessels and take advantage of the situation. 

The bactericidal state, therefore, enfeebles the viru- 
lent microbes, which are then overcome and removed 
by the phagocytes. 

Immunity being in some way fixed in the anatomi- 
cal elements, one can understand its persistence and 
the possibility of its hereditary transmission. In 
natural immunity or the refractory condition the 
bactericidal property does not appear to have, at 
least in certain cases, the same importance as in ac- 
quired immunity. Indeed, the blood of an individual 
refractory to a given microbe can serve perfectly for 
the artificial culture of this microbe. Nevertheless, 
too much importance should not be attached to this 
fact. We shall see, later, that the blood is always 
more or less bactericidal, but that this faculty disap- 
pears shortly after its removal from the vessels. The 
bactericidal property of the blood, therefore, depends, 
without doubt, upon its contact with the living tis- 
sues, hence we can form no accurate estimate of this 
condition of the blood contained in the vessels of the 
organism by comparing it with the same fluid outside 
of the vessels. 

"We know, again, that natural immunity results, in 
the case of certain microbes, on account of the tem- 
perature of the organism being either too high or too 
low; it may also, as M. Arloing has demonstrated, 
be the result of a natural insusceptibility of the or- 
ganism to the action of the amorphous products se- 
creted by the microbes. 



74 Manual of Veterinary Microbiology. 



II. Reaction of the organism against microbes. 

We will consider, successively, under this head: 
phagocytosis, the bactericidal property, the isolation 
and elimination of pathogenic microbes, and varia- 
tions of virulence produced by the organism. 

When non-pathogenic germs, or germs in which 
the virulence is extinguished, are introduced into the 
tissues of an animal, these germs are more or less 
quickly destroyed. The same thing occurs when 
pathogenic germs in full virulence are introduced 
into the system of an individual destitute of recep- 
tivity for the germs. On the other hand, if the viru- 
lent germs are brought into contact with an organism 
endowed with receptivity they will multiply and be- 
come the starting point of morbid troubles. 

There are, therefore, in the organism of certain ani- 
mals, conditions capable of bringing about the d'e- 
struction of microbes. These conditions are multiple 
and their nature is as yet incompletely elucidated. 
We know, however, that the organism defends itself 
against invasion through the intermediation of its 
figured elements and of its fluid parts. 

1. Phagocytosis. The name phagocytosis is given to 
the destructive action of certain cells toward mi- 
crobes, these cells being known as phagocytes. The 
white corpuscles, in this regard, take first place in 
whatever part of the body we find them : blood, con- 
nective tissue, lymphoid organs (spleen, lyrnph glands, 
marrow of bones, etc.) ; then come the fixed cells of 
the connective tissue, the endothelial cells of the 
capillary vessels, the cells of soft epithelia, muscular 
fibers, etc. 



Physiology of Pathogenic Microbes. 75 

The disease develops when the phagocytes do not 
succeed in killing the introduced germs ; in the op- 
posite case it fails to develop. 

The injection of virulent germs in a susceptible in- 
dividual paratyzes the phagocytes ; on the contrary, 
the injection of attenuated germs is followed by an 
accumulation of leucocytes (diapedesis) around the 
place where the germs occur; the germs are then 
taken into the substance 'of the leucocytes and di- 
gested by them. The same thing happens when we 
inject virulent germs into a non-susceptible subject. 

All the germs seized by the phagocytes are not in- 
fallibly destroyed; hence, these migratory cells, in 
certain cases, seem to be a means of transferring 
bacteria from one part of the organism to another. 

The leucocytes possess, in common with some of 
the lower vegetables, the property (called chemotaxis) 
of moving themselves toward certain chemical sub; 
stances. Now, experiments have shown that the 
sterilized or filtered cultures of a certain number of 
microbes attract the white corpuscles, whilst cultures 
of other microbes either have no action upon white 
corpuscles or paralyze their movements. 

Some germs secrete substances which paralyze the 
vaso-dilator nerves, thus opposing a direct obstacle to 
diapedesis and therefore to phagocytosis. As an il- 
lustration of this nervous action we find that the ear 
of the rabbit which has received an injection of the 
soluble products of the pyocyanic bacillus, for ex- 
ample, does not become inflamed when a layer of 
croton oil is applied to it. 



70 Manual of Veterinary Microbiology. 

Microbes whose secretions attract the leucocytes * 
will be more easily overcome by them, and will be 
less dangerous for the economy. 

Microbes which repel the white corpuscles, finding 
themselves in good conditions as to nutrition and 
surroundings, will break down the barrier which the 
corpuscles oppose to them. But, under conditions 
unfavorable to their development, their secretions 
diminish and the phagocytes regain all their power. 

Some agents, such as chloral and chloroform, are 
capable of suspending the chemotaxic faculty of the 
phagocytes. 

Certain influences physical and moral disturb- 
ances, fatigue, nervous perturbation, cold, which often 
cause the irruption of an infectious disease or aggra- 
vate it have a depressing effect upon the vaso-dilator 
nervous apparatus, interfere with diapedesis, with 
phagocytosis, and therefore favor the implantation or 
multiplication of the germs of disease. 

2. Bactericidal or microbicidal property. By the bac- 
tericidal property is meant the peculiar quality of the 
humors of the economy blood, aqueous humor, peri- 
cardial serosity, etc. which impedes or prevents the 
multiplication of pathogenic bacteria in these fluids. 
This bactericidal faculty varies greatly according to 
the species, the individual, and the germs with which 
we have to do. When microbes are introduced into 
the blood, a certain number of them perish ; those 

* [According to Buchner, the attractive action (positive chemo- 
taxis) exerted by sterilized cultures of certain microbes toward 
leucocytes is dependent on the proteid contents of the bacterial 
cells, rather than on their secretion products. Baumgarten's 
Jahresbericht, 1890. D.] 



Physiology of Pathogenic Microbes. 77 

which resist are then capable of multiplying ; hence, 
the normal microbicidal property of the blood is tem- 
porary, not permanent. Fresh blood kills the bacillus 
of charbon, but constitutes a suitable medium for its 
cultivation eight days after its removal from the ves- 
sels. It seems to be established that, when we inject 
these bacilli into the blood, they multiply there only 
after having previously multiplied in one or more 
organs in which they have been arrested. When 
only a small number of microbes have been intro- 
duced they may all be destroyed, and then the inocu- 
lation fails. 

Non-pathogenic microbes, introduced into the 
blood, disappear from this fluid, becoming arrested 
in the fine capillaries of the liver, spleen, marrow of 
bones, and kidneys, in which situation they are 
quickly destroyed. 

Pathogenic microbes have a similar experience 
when they are inoculated in small doses into the cir- 
culation ; like the preceding, they are consigned to 
the fine capillaries of the parenchymatous organs 
and there sustain the conflict with the phagocytic 
elements (endothelial cells, white corpuscles, etc.) 
According as the issue of this conflict proves favor- 
able to the microbes or to the phagocytes we may 
expect the appearance or non-appearance of the dis- 
ease. 

The evolution of the microbic disease is accom- 
panied, when the disease is non-recurrent, by the pro- 
duction of the bactericidal property. This condition 
is slowly evolved by the progressive action of the 
soluble microbic products upon the fluids and tissues. 
When it has acquired sufficient intensity it may lead 



78 Manual of Veterinary Microbiology. . 

to the limitation of, or recovery from, the disease, by 
opposing itself to the multiplication, 'and interfering 
with the nutrition of the introduced germs ; the 
phagocytes then assume the duty of freeing the or- 
ganism from these enfeebled germs. 

Elimination of microbes. The infected organism 
may free itself from microbes by the local reaction 
which their presence excites. The germs of pus, for 
example, determine around them the diapedesis of 
leucocytes and these destroy by phagocytosis a large 
number of germs. On the other hand, at the same 
time that the accumulation of leucocytes within the 
meshes of the vascular network presents a certain 
degree of obstruction to the nutrition of the mi- 
crobes, the microbic secretions act upon the leuco- 
cytes, either by virtue of their dissolving disastases or 
of their toxic products, so that a parallel destruction 
ensues of white corpuscles and of microbes. Whilst 
this double destruction goes on at the center of the 
focus, the surviving germs continue to invade the 
peripheral tissues until there is formed all around the 
invading army a sufficient barrier of phagocytes. 
We then see the limitation of the abscess, and, occa- 
sionally, its encystment by fibrous organization of 
the limiting tissues ; but more frequently the exten- 
sion of the pyogenic inflammation gives rise to soften- 
ing, perforation of the integument, and the elimina- 
tion of the pus and therefore of the germs, the cause 
of all the trouble. 

The natural elimination of pathogenic microbes can 
occur in all cases in which the lesions they deter- 
mine are situated near the surface of the skin or 
mucous membranes communicating directly with the 



Physiology of Pathogenic Microbes. 79 

exterior ; this elimination is more or less complete' 
according to the case. 

When the germs are deeply situated their expulsion 
may still take place through certain glands the kid- 
neys, the salivary glands (rabies), the liver (symptom- 
atic charbon). The elimination of infectious germs 
by way of the kidneys has been observed for a large 
number of diseases; generally, if not always, it is 
the consequence of the irritation which the microbes 
or their products exert upon the organ. Many mi- 
crobic diseases, in fact, are accompanied by nephritis. 
The more or less complete desquamation of the 
secretory epithelium, and the vascular troubles which 
then supervene account for the passage of the mi- 
crobes. 

If the elimination of the figured agents of the in- 
fectious diseases demands conditions somewhat com- 
plex, the rejection of their secretions is more readily 
accomplished, since these substances are dissolved in 
the liquid media of the economy. Their filtration 
takes place especially through the kidneys; their 
presence in the urine has been established; but they 
may also transude from the various other natural 
emunctories. 

4. Modifications of virulence The bactericidal prop- 
erty of the humors has the power of attenuating the 
virulence of pathogenic microbes. Inversely, the 
absence of this property may occasion an increase 
of this virulence. Thus, the virulence of a given 
microbe may become increased or diminished by 
passing through a series of individuals of the same 
species : rouget of the pig becomes more active when 
it is made to pass through the pigeon or the rabbit ; 



80 Manual of Veterinary Microbiology. 

on the contrary, the organism of the ape attenuates 
the virus of rabies. 

The virulence, modified by one species,, may be 
changed in the same or opposite direction for other 
species : rouget which has become more virulent in 
the rabbit is less virulent for the pig; whilst the pass- 
age of the same germ through the pigeon augments 
its pathogenic power not only for the pigeon but also 
for the pig. 

We could multiply examples of the influence of 
the natural organic media upon pathogenic germs. 
The virus of rabies attenuated in the ape is also at- 
tenuated for the dog, guinea pig, and rabbit. Inocu- 
lated in series to one of these species, it regains the 
virulence which it naturally possesses in the dog; in 
the guinea pig and rabbit it may even surpass this, 
and in the latter attain a maximum of activity be 
yond which further passages no more modify it. 

III. Evolution of the bacterial disease. 

After what we have seen of the reciprocal action 
of the organism arid pathogenic microbes, little re- 
mains to be said of the disease itself. 

The determining cause of microbic affections re- 
sides always in the implantation of the specific germ 
in a susceptible individual ; but such common causes 
as cold, mental emotions, overwork, etc., may take 
an important part in their etiology, by diminishing 
or suspending the normal phagocytic action, as we 
have already seen. 

The germs once introduced and having resisted the 
combined influence of phagocytosis and the bacteri- 
cidal property, their effects do not immediately become 



Physiology of Pathogenic Microbes. 81 

appreciable. A preparatory period then ensues dur- 
ing which the microbe proliferates, multiplying its 
means of action upon the economy. This period 
corresponds to the incubation. It is shorter the bet- 
ter the organism is adapted to the life of the germ; 
its duration also varies with the nature of the germ 
and its virulence, with the abundance of the virus, 
with the receptivity of the subject, the place of inoc- 
ulation, etc. In some diseases the duration of the 
incubatory period is almost constant; in others it is 
very variable (rabies). 

Certain diseases have several successive incuba- 
tions, or, rather, latent periods, during which the dis- 
ease germ ceases its activity, slumbers, to resume at 
a later period its course of invasion (tuberculosis, 
dourine, syphilis). The organism is then in the con- 
dition of latent microbism. 

The period of incubation is completed when the 
first manifestations of the morbific action of the virus 
appear; the premonitary symptoms are in no way 
characteristic ; yet, in the course of an epizootic, 
there are certain signs by which we can recognize 
the invasion of the disease in an individual previously 
healthy. Thus, a persistent fever in one of the cattle 
of a stable where pleuro-pneumonia prevails, would 
excite a suspicion of its invasion by this disease. 

The early obscure symptoms more or less quickly 
give place to troubles more and more serious, which 
clearly characterize the affection with which we have 
to do, and express the influence of the progressive 
action of the microbe. This is the period of increase. 
We will not enter here into details of the common 
symptoms of microbic diseases, an outline of which 



82 Manual of Veterinary Microbiology. 

has been incidentally given in connection with the 
subject of their pathogeny. We will only say a few 
words on the specificity of pathogenic microbes. At 
an early period in bacteriological research investi- 
gators applied themselves to the discovery of a spe- 
cial microbe for each disease; it was believed that 
every germ produced always the same effects. We 
now know, however, that the same microbe may give 
rise to very different diseases : the microbe of fowl 
cholera gives a true septicaemia in the rabbit, a cir- 
cumscribed abscess in the guinea pig; the streptococ- 
cus pyogenes develops sometimes an abscess, some- 
times an erysipelas, sometimes puerperal fever. Sim- 
ilarly, a given lesion may be consecutive to the inoc- 
ulation of different microbes: the particular inflam- 
matory lesion which has received the name of tuber- 
cle has for its usual cause the tubercle bacillus; but 
other agents can develop identical changes, which 
have been designated pseudo-tubercles in order to dis- 
tinguish them from those in which the said bacillus 
exists; among the number of these agents we may 
mention croton-oil, lycopodium spores, the demodex 
folliculorum, strongylus vasorum, the utricular sarco- 
spermia of the muscles of the pig, the actinomyces, 
the pseudo-tubercle bacillus of Courmont, etc. Ty- 
phoid fever of the horse appears to develop under the 
influence of various species of microbes, acting inde- 
pendently. 

Microbic specificity is, therefore, not absolute; it 
depends upon the organism in which the parasite 
has implanted itself and on the external conditions 
which may have influenced the latter. Attenuation 
of a microbe suffices, indeed, to change its effects. 



Physiology of Pathogenic Microbes. 83 

An infectious disease may be local or general, ac- 
cording as the germs are confined to the place of in- 
oculation or have invaded the circulation. In the lat- 
ter case the general affection may have been pre- 
ceded by circumscribed local lesions, or it may 
have been generalized from the first. In either case, 
a general disease can determine localized lesions, 
specific or not : nephritis, hepatitis, enteritis, inflam- 
matory enlargements, etc. These secondary inflam- 
mations of the secretory organs may be the starting 
point of grave complications (auto -intoxications): ab- 
sorption of bile, urinary intoxication. Changes of 
these organs present, in addition, direct obstacles 
to the elimination of the soluble microbic products. 

The multiplication of disease germs arrives, at the 
end of a certain time, at its apogee ; the disease then 
reaches its height. The* secretion-products of the 
microbes become harmful to the microbes themselves ; 
in mingling with the fluids they communicate to 
them, as well as to the tissues, the faculty of check- 
ing microbic proliferation, the microbicidal faculty ; 
in a word, they vaccinate the organism. The vac- 
cinating effect is slowly produced under the influence 
of the prolonged contact of these products. Im- 
poverishment of the organic media in principles in- 
dispensable to the microbes, febrile elevation of tem- 
perature, can also act prejudicially upon the latter. 
From this ensemble of unfavorable circumstances 
there results enfeeblement of the germs, and the 
phagocytes take upon themselves the task of destroy- 
ing them. 

The disease then subsides on account of the fact 
that the toxic substances of microbic origin which 



84 Manual of Veterinary Microbiology. 

become eliminated in various ways, are no longer re- 
placed by fresh additions. Recovery is not complete 
when the elimination of these substances is ended. 
Besides the weakness of the patient, local troubles 
more or less important may persist, recovery from 
which will take place gradually now that the cause 
which engendered them is removed. 

But the disease, having attained its height, may 
terminate in death, the manner in which this is 
brought about varying in different cases. 

Microbicdiseas.es maybe acute or chronic, epizootic, en- 
zootic or sporadic. The gravity of some, at least, of 
these diseases is subject to variations, whilst others of 
them are almost invariably fatal : charbon and rabies 
for example. Indeed, from the knowledge which we 
have gained in regard to receptivity, immunity, and 
the resistance of the organism we should expect all 
degrees of intensity in such diseases. In epizootics of 
great severity it is observed that the first animals at- 
tacked are more severely affected than the last, and 
that the number of the individuals attacked, consider- 
able at the beginning, rapidly diminishes toward the 
end of the attack. This is probably due to the fact 
that the virus fixes itself upon individual animals by 
reason of their special susceptibility. The least refrac- 
tory will be first attacked, will be severely affected, 
and the virus will rapidly multiply, thus increasing 
the chances of infection for all those which are sus- 
ceptible to the disease. On account of this dissemi- 
nation of the virulent germs the less susceptible will 
also be finally stricken, but, by reason of the fact that 
they do not offer a very favorable field, the microbic 
pullulation will in these be less extensive, the disease 



Transformation, etc., of Pathogenic Microbes. 85 

less serious, and the chances of infection for the indi- 
viduals yet exempt and which are, moreover, the 
most refractory, will rapidly diminish. 

The disease may be continuous (charbon), remittent^ 
(tuberculosis), or intermittent (malaria). 



CHAPTER III. 

TRANSFORMATION AND DESTRUCTION OF PATHOGENIC MI- 
CROBES IN THEIR RELATION WITH HYGIENE AND 
THERAPEUTICS. 

1. Morphological and physiological variations of pathogenic mi- 
crobes. 2. Attenuation. 3. Preventive inoculations. Vac- 
cinations. 4. Destruction and annihilation of pathogenic 
germs. 

1. Morphological and physiological variations of patho- 
genic microbes. 

We have already, in the first part of this work, re- 
ferred briefly to the influence of the media upon mi- 
crobes in general. This influence acquires great 
importance when pathogenic germs are in question. 
The latter may undergo considerable changes under 
the influence of external conditions. 

Pathogenic germs undergo, from this cause, varia- 
tions of form and of function. 

Physical variations are quite common : the microbe 
of avian cholera shrinks when its cultures become 
old; that of rouget, cultivated after its passage 



86 Manual of Veterinary Microbiology. 

through the rabbit, increases in volume; the bacillus 
of the pyocyanic disease presents itself, according 
to the media in which it is cultivated, as a bacillus, 
a spirillum, or a micrococcus; the bacillus of gan- 
grenous septicaemia grows in the form of short rods 
in the connective tissue, in long filaments in the 
serous membranes and blood. The bacteridium of 
charbon, cultivated in bouillon containing a small 
quantity of bichromate of potassium, loses its power 
of forming spores, subsequent generations to which 
it gives birth being also asporogenous. A tempera- 
ture of 42 to 43 produces the same effect upon the 
bacteridium. 

The functional variations are of more importance ; 
the pyocyanic bacillus, placed under certain condi- 
tions, ceases to secrete the coloring matter which 
characterizes it. There are, however, variations of 
virulence which more especially interest us. The 
virulence of pathogenic germs may be increased or 
diminished, then brought back to its normal intensity, 
by conditions which vary with each microbe; these 
conditions will be considered more in detail in the 
following paragraph. 

The virulence of a microbe may become enfeebled 
to such an extent as to completely lose its pathogenic 
powers ; it then becomes saprogenic. From the rec- 
ognition of this fact to the admission of microbic 
transformation there is only a step. Nevertheless, 
up to the present, we have not observed the forma- 
tion of a new species at the expense of another 
species. From a practical point of view, however, it 
must be admitted that certain saprogenic species may 



Transformation, etc., of Pathogenic Microbes. 87 

incidentally become pathogenic and determine the 
development of morbid troubles in man or animals. 

II. Attenuation of pathogenic microbes. 

The diminution of virulence of pathogenic microbes 
is occasionally, not always, connected with a diminu- 
tion in their general nutritive activity ; it may occur 
under quite varied conditions, either spontaneously 
or from certain definite influences purposely brought 
into play by the experimenter; we will here study 
the different means by which attenuation may be ob- 
tained. 

1. Attenuation by the normal atmosphere. We al- 
ready know that the majority of the atmospheric 
germs are dead. It is logical to admit that the loss 
of their vitality did not take place abruptly, but, on 
the contrary, was gradually produced, and that what 
virulence they may originally have possessed also dis- 
appeared gradually. In short, the germs of the air 
are attenuated before being destroyed. The atmos- 
pheric conditions which determine these changes in 
microbic life are far from being simple : oxygen, 
light, the electrical condition, desiccation, probably 
all act in concert. 

But the external air, of itself alone, can bring 
about the attenuation of pathogenic germs. Very 
active cultures may lose their virulence in some days, 
weeks, or occasionally months. 

Cultures of fowl cholera, abandoned to the air, 
gradually diminish in virulence so as to completely 
lose it at the end of a time varying from six weeks 
to two months, occasionally much less. By re-sowing 
these germs, in way of attenuation, at periods more 



88 Manual of Veterinary Microbiology. 

and more remote from the establishment of the cul- 
ture, we obtain generations of progressively decreas- 
ing virulence, a series of viruses less and less power- 
ful, the special activity of which is preserved when 
we exclude them from contact with the air or when 
we rejuvenate them without intermission by cultures 
made at very short intervals. Attenuation is here, 
therefore, hereditary through successive generations. 
Those of most feeble virulence constitute vaccines 
against more virulent cultures. 

Cultures of the germ of rouget undergo changes 
similar to those of fowl cholera. 

As to cultures of the charbon bacillus, they are 
infinitely more resisting to the destructive influence, 
of the atmosphere. This is because they contain 
spores, and when we wish to attenuate them in con- 
tact with the air it is necessary to begin by prevent- 
ing sporulation. Pasteur has attained this end by 
cultivating charbon at a temperature of 42 to 43. 
At this temperature, the culture exposed to contact 
with the air, rapidly loses its virulence ; it ceases to be 
fatal first for the larger animals, then for small adults, 
finally, for small animals only a few days old.. The 
bacteridium itself perishes much more slowly. Now, 
each degree of virulence can be perpetuated by cul- 
tivating at 42 to 43 the -different varieties obtained, 
each of these varieties transmitting its special viru- 
lence to its descendants. If they are returned to 87 
they form spores possessing in embryo the special 
pathogenic activity of the attenuated bacilli from 
which they originate, and susceptible of transmitting 
the latter to new generations cultivated at 37. 

According to M. Chauveau,the attenuation of the 



Transformation, etc., of Pathogenic Microbes. 89 

bacteridia by the preceding process is the effect of 
the heat and not of the oxygen, as was advanced by 
Pasteur. 

The least virulent varieties produce a mild disease 
which leaves behind it immunity for the varieties less 
attenuated, and we thus have vaccines of different 
degrees of strength which we can bring into action 
in succession. 

The attenuated virus of fowl cholera and of bac- 
teridian charbon can be restored to their normal 
virulence by passing them through the organism, 
first, of young individuals, and then through those 
of individuals of gradually increasing age. 

2. Attenuation by compressed oxygen or air. M. 
Chauveau has succeeded in attenuating the bacteridium 
of charbon by subjecting it to contact with pure oxy- 
gen at a pressure of two and a half atmospheres dur- 
ing fifteen days and over, at a temperature of 35 to 
36. This attenuation is transmitted through suc- 
cessive generations of the bacteridia. He has, in 
this way obtained an energetic vaccine which con- 
fers immunity against charbon without giving rise to 
accidents in the vaccinated, the vaccinating property 
being wholly retained whilst the virulence becomes 
progressively enfeebled until it entirely disappears. 

3. Attenuation by heat. Heat is a powerful means 
of destruction of germs; we also find it among the 
number of attenuating agents. It is to Toussaint 
that the honor belongs of having first demonstrated 
this property of heat in connection with charbonous 
blood. His methods were improved, and Chauveau 
applied himself to the task of heating small quanti- 

8 



90 Manual of Veterinary Microbiology. 

ties of charbonous blood in such a manner that all the 
bacilli were influenced to the same extent; the de- 
gree of enfeeblement, in fact, depends on the temper- 
ature and the duration of the heating. Chauveau has 
prepared two vaccines, intended to be inoculated in 
succession : the first, the least active, is obtained by 
heating defibrinated blood at 50 during fifteen min- 
utes ; the second, by heating the same liquid at the 
same temperature during nine to ten minutes. This 
last vaccine inoculated to animals which have not re- 
ceived the first may yet cause the death of some in- 
dividuals. 

To obtain a uniform result with charbonous blood 
it is necessary always to use fresh blood in which the 
bacilli are free from spores. These latter are endowed 
with a much greater power of resistance and are not 
attenuated like the bacilli; hence, in his researches 
upon the attenuation of cultures, M. Chauveau first 
cultivated the bacteridia at a temperature of 42 to 
43, in order to prevent the formation of spores. The 
asporogenous bacteridia are heated to 47 during three 
consecutive hours ; they are then attenuated to the 
extent that they no longer kill adult guinea pigs. 
Brought then to the favorable vegetating tempera- 
ture of 37 the attenuated bacteridia form spores ; but 
these are liable when placed under suitable conditions 
to produce virulent bacilli again. 

In order to render attenuation transmissible to suc- 
cessive generations of bacteridia M. Chauveau then 
brought the sporulated cultures to temperatures 
neighboring on 80. The attenuation then became 
fixed upon the spores and the latter, returned into 
nutritive bouillon at 37, produces bacilli attenuated 



Transformation, etc., of Pathogenic Microbes. 91 

like themselves. Of two vaccines prepared in this 
way, the first comes from heating at 84 during one 
hour, the second from heating at 82 during the same 
time. 

The natural serosity of the specific lesions of symp- 
tomatic charbon may also be attenuated to different 
degrees by a temperature of 65 to 70, maintained 
for a greater or less period of time (Arloing, Corne- 
vin and Thomas). However, these authors work in 
preference with serosity dried at a temperature of 30 
to 35 ; the dried virus is, in fact, more fixed than the 
fluid serosity ; it can be preserved indefinitely with its 
normal virulence; its attenuation requires tempera- 
tures varying between 60 and 110. The authors 
have prepared two vaccines from it: one heated at 
100, the other at 85; Kitt has recommended a single 
vaccine heated at 90. 

4. Attenuation by solar light. Light is a powerful 
bactericidal agent ; by the careful use of this prop- 
erty M. Arloing has succeeded in gradually attenua- 
ting the charbon bacillus. 

" Thus, a culture exposed to the rays of the sun 
during nineteen hours furnishes a virus which kills 
the guinea pig in the do.se of one drop ; exposed dur- 
ing twenty hours, a culture only kills one guinea pig 
out of two ; exposed during twenty-five hours, such 
a culture no more kills guinea pigs but vaccinates 
them, the vegetating power of the bacillus being also 
considerably diminished." 

The diminution of virulence thus obtained is tem- 
porary and is not transmitted to later generations of 
the bacilli. 

5. Desiccation. Desiccation has been utilized by M. 



92 Manual of Veterinary Microbiology. 

Pasteur in attenuating the virus of rabies. The cords 
of rabid rabbits, suspended in vessels containing 
pieces of caustic potash and maintained at a tempera- 
ture of 20, lose their virulence in seven days; the 
diminution takes place progressively, so that, by fix- 
ing the successive degrees of virulence by inocula- 
tion to the rabbit, we can obtain virus of gradually 
increasing intensity. 

Attenuation is here the result of several factors : 
desiccation, oxygen, and the temperature. The in- 
fluence of the last agent is itself very great if we 
can judge by the increased rapidity with which at- 
tenuation is produced when the temperature is even 
slightly increased ; the virulence is, in fact, obliterated 
in seven days at 20, in five days at 23, and in 24 
hours at 35. 

6. Attenuation by antiseptics. Chemical substances 
which are toxic for bacteria can diminish the vitality 
of the latter when they are employed in selected doses 
and their contact maintained during definite periods 
of time. 

The bacillus of symptomatic charbon is attenuated 
to the point of becoming a vaccine in contact with 
sublimate, at 1 to 5000, with carbolated glycerine, 
eucalyptol, and thymol. Carbolic acid, at 1 to 800, 
allows the multiplication of the charbon bacteridium 
but prevents sporulation. Continuation of the con- 
tact with this solution gradually weakens the viru- 
lence. The same results are obtained, but much 
more rapidly, with bichromate of potassium in the 
proportion of from 1 in 2000 to 1 in 5000. 

The soluble substances secreted by germs can di- 
minish the virulence of other germs ;' thus bouillon 



Transformation, etc., of Pathogenic Microbes. 98 

containing the residues of a culture of cholera atten- 
uates the charbon bacillus. 

7. Attenuation by passage through the organism of 
animals. Pathogenic germs are subjected, by the or- 
ganisms upon which they implant themselves, to cer- 
tain influences of which we have already had occa- 
sion to speak; we have noted especially the modifi- 
cations which may supervene in the virulence of these 
germs either in the way of increase or diminution. 

Instances of attenuation being produced under 
these conditions are quite numerous. 

The microbe of rouget of the pig becomes well ac- 
climated in the pigeon and the rabbit, and in these 
two species acquires great virulence ; but, whilst the 
organism of the pigeon renders it more active for the 
pig, that of the rabbit diminishes its virulence for 
this animal. This attenuation is preserved in cul- 
tures then made in ordinary bouillon, and these cul- 
tures can be employed as vaccine for the pig. 

The bacilli of bacteridian charbon and of symp- 
tomatic charbon are attenuated by their passage 
within the lymphatic sacs of the frog. 

The virus of rabies is attenuated in passing through 
the organism of the ape to the exte-nt of. becoming 
inoffensive for the dog and of vaccinating it. It re- 
sults from the preceding facts that the morbigenic 
faculty of microbes can be lessened in different de- 
grees ; in certain cases this diminution only exists for 
those bacteria upon which the conditions determin- 
ing the attenuation have acted, and the attenuation 
is temporary ; in others the modification to which the 
microbes have been subjected is more profound, more 



94 Manual of Veterinary Microbiology. 

durable, and persists in later generations of these 
microbes. 

The methods which furnish an hereditary attenu- 
ation permit of obtaining more easily large quantities 
of attenuated virus, and are more especially utilized 
in the preparation of vaccines. 

III. Preventive inoculations. Vaccinations. 

The organism may be made refractory to a bac- 
terial disease by different means ^ by inoculating the 
natural virus, the attenuated virus, a chemical vaccine, 
or, finally, a virus different from that against which 
it is desired to fortify the organism. 

1. Preventive inoculation of natural virus. It has 
been observed that a certain number of -contagious 
diseases leave behind them in the subject, after re- 
covery, a solid immunity against these same diseases. 
On the other hand, certain diseases purposely com- 
municated show themselves much less dangerous than 
when they prevail naturally. The recognition of 
these facts has given origin to variolization, clavel- 
ization,* preventive inoculation against pleuro-pneu- 
monia, etc. The first, for more than a century, has 
been replaced by vaccination; the second is still 
recommended in our times, in default of a means of 
prevention more practical, if not more efficacious. 

In this case the only object is to produce a disease 
of the same nature as the spontaneous disease, but 
mild in character, not threatening the life of the in- 
dividual, yet endowing it with immunity. This end 
is obtained by diminishing the number of the germs 

* [Artificial infection of flocks with ovine variola. D.] 



Transformation, etc., of Pathogenic Microbes. 95 

which engender the disease; the organism overcomes a 
small dose of a certain virus when it would be over- 
come by a larger dose : a minimum quantity of the 
virus of gangrenous septicaemia and of symptomatic 
charbon vaccinates against these diseases; a larger 
quantity produces the fatal disease. Dilution of the 
vaccine, of the virus of sheep-pox, lessens its effects. 
The severity of a disease may, again, be diminished 
by introducing the virus by a special way known to 
mitigate its influence : the blood (pleuro -pneumonia, 
symptomatic charbon, gangrenous septicaemia, rabies), 
the cellular tissue of the tail (pleuro-pneumonia). 

2. Preventive inoculation of attenuated virus. Atten- 
uated viruses develop a mild disease which confers on 
the animals -an immunity more solid the less the de- 
gree of attenuation ; in general, we have recourse in 
practice to several specimens of virus of different de- 
grees of intensity ; we begin with the weakest and 
end with the strongest; a solid immunity may thus 
be communicated by virus sometimes yet very active 
but against the action of which the less virulent va- 
rieties have fortified the organism. 

Attenuated viruses which are able to prevent the 
development of infectious diseases are called vaccine 
viruses or simply vaccines. 

The two charbons, rouget, pneumo-enteritis of the 
pig, chicken cholera, rabies, etc., have furnished vac- 
cines the employment of which is to-day admitted 
into general practice. 

3. Preventive inoculation of soluble vaccinating sub- 
stances. We have seen above, in connection with the 
subject of acquired immunity, that the latter results 
from the impregnation of the organism with the solu- 






96 Manual of Veterinary Microbiology. 

ble products produced by pathogenic bacteria. These 
vaccinating substances are known for a certain num- 
ber of diseases the pyocyanic disease, bacteridian 
charbon and symptomatic charbon, cholera, Pasteur's 
septicaemia, pneumo-enteritis of the pig, rabies, hu- 
man tuberculosis (Koch's lymph will give immunity 
to guinea pigs), tuberculosis of birds (Courmont and 
Dor have vaccinated the rabbit by means of the solu- 
ble products of cultures), etc. 

These substances, also called chemical vaccines, 
have a great advantage over the figured elements; 
the attenuated virus may indeed, in exceptional cases, 
regain its virulent properties, and this unknown to 
the experimenter; it then produces the fatal disease 
instead of the immunity which was expected of it. 
The soluble vaccinating substances, however, are not 
entirely free from danger; they are nearly always 
extremely violent poisons, and the quantities em- 
ployed must be judiciously regulated. The discovery 
of these substances is of such recent date that in 
practice we have not as yet reaped the benefits from 
them which we may reasonably expect. 

Attempts at the prevention and cure of the mi- 
crobic diseases by organic liquids coming from species 
naturally refractory to these diseases have been made 
in recent times ; here, again, the action concerned is 
a chemical one. The blood of the goat, transfused 
to the rabbit at the time that the latter is inoculated 
with tubercular products, will prevent the evolution 
of tuberculosis; if the transfusion is made after the 
disease has already commenced, it will retard it and 
may even cause its retrogression. 

Injection of the blood serum of^the dog (haemocyne) 



Transformation, etc., of Pathogenic Microbes. 97 

vaccinates the rabbit against tuberculosis ; this pre- 
servative action is more intense with the serum of a 
tuberculized dog and manifests itself even when the 
injection is made seven days after the virulent inocu- 
lation. 

We must mention here, in connection with pre- 
ventives of a chemical nature, the attempts at vac- 
cination against rabies with the essence of tansy, 
against tetanus with strychnine, and against tubercu- 
losis with tannin. 

4. Preventive inoculation with the virus of another 
disease. Cow-pox is preservative against variola; we 
place the example in this paragraph although, ac- 
cording to recent researches, it should have its place 
along with the inoculations of attenuated virus. Re- 
cent experiments of M. Eternod have, in fact, demon- 
strated the identity of variola with vaccinia. It 
should be said, however, that these results have been 
contested. 

The attenuated microbe of fowl cholera vaccinates 
fowls against charbon and against Davaine's septi- 
caemia; guinea pigs vaccinated against symptomatic 
charbon are also vaccinated against the septic vibrio, 
but the reverse does not hold good. 

IV. Destruction and annihilation of pathogenic germs. 
The destruction of pathogenic germs is a very im- 
portant point to consider. The hygienist and the 
physician should apply themselves to the suppression 
of dangerous germs wherever they exist, that is, 
upon the patient and upon the objects which have 
been contaminated in his surroundings; this opera- 
9 



98 Manual of Veterinary Microbiology. 

tion is known as disinfection. Nature comes power- 
fully to their aid in this work of purification ; but it 
often requires to be seconded by artificial means, the 
application of which generally abridges the natural 
duration of pandemic or panzootic diseases. We 
have therefore to separate the causes which bring 
about the destruction of pathogenic germs into 
natural and artificial. 

It may be stated as a fundamental principle that 
adult bacteria in their vegetative form are more rap- 
idly destroyed, whatever be the cause of destruction, 
than the spores or organs of fructification. For the 
vegetative forms the action of destructive agents is 
more or less rapid according to the case ; a bacterium, 
taken in full vegetation in a suitable medium, will be 
more easily killed than one in the condition of latent 
life, for example, in a state of desiccation, which is 
equivalent to saying that death will be more easily 
produced as life is more intense, more complete. 
Something of the same kind is observed in higher 
beings, the liability of these to suffer from adverse 
conditions being greater as their requirements are 
the more exacting. A young bacterium recently de- 
veloped from a spore is more sensitive than an adult 
bacterium which has attained its complete develop- 
ment. 

1. Natural disinfecting agents. These are light, des- 
iccation, and oxygen. 

Light. Light excites oxidation of the organic sub- 
stances which enter into the constitution of germs, 
and, as a consequence, involves the death of the lat- 
ter. Other conditions being the same, solar light 
acts more rapidly upon non-sporulated germs than 



Transformation, etc., of Pathogenic Microbes. 99 

upon spores, upon germs contained in a liquid or 
moist medium than upon those which are in a dry 
medium. 

The time necessary for destruction by light varies 
from a few hours to some weeks, according to the 
case ; solar light, however, is a sure agent and one 
whose beneficial action operates in a continuous man- 
ner ; hence, we should guard against voluntarily de- 
priving ourselves of it ; abundance of light in inhab- 
ited places is one of the most rational of hygienic 
measures. 

Desiccation. Insufficiency of water arrests the mul- 
tiplication of microbes ; the latter then lose their vi- 
tality more or less rapidly. But the spores resist 
much longer than the bacteria themselves ; we know, 
indeed, that the virus of symptomatic charbon is 
dried in order to preserve it, and that the germs of 
tuberculosis are preserved active for a long time in 
pulverulent sputum. 

Oxygen. The germs of the air are destroyed more 
or less quickly under the combined influence of des- 
iccation, which arrests their pullulation and impairs 
their vitality, of oxygen, which oxidizes them, and 
of the solar light, which excites in them this oxida- 
tion; the oxidizing action of oxygen, without doubt, 
extends itself to the bacteria of waters and of the 
soil, these perishing more or less rapidly by reason of 
the conditions unfavorable to their multiplication 
which they meet in these media. 

Other natural agents, less important, may intervene 
to destroy pathogenic bacteria; among these we will 
notice again the ordinary saprogenic germs which, 
finding themselves in the same media as the patho- 



100 Manual of Veterinary Microbiology. 

genie, may lead directly to the death of the latter; 
we have already explained this action. 

2. Artificial disinfecting agents. These are heat and 
the antiseptics. 

Heat. Heat is a powerful means of disinfection. 
Although germs withstand extremely low tempera- 
tures, they are very sensitive to increase of temper- 
ature. Temperatures too low, as well as those which 
are too high, are unfavorable to the proliferation of 
pathogenic microbes; but the latter are, in addition, 
rapidly fatal. Nevertheless, the spores are infinitely 
more resistant than the adult germs; whilst these 
last are killed in all cases between 50 and 100, the 
spores, on the contrary, require a temperature of 
110 to 125. However, we can with certainty kill all 
spores in a given infusion by bringing, on different 
occasions, and at intervals of one day, the said in- 
fusion to a temperature fatal for the adult form, and 
taking the precaution between the periods of heat- 
ing to place the whole at a temperature favorable 
for vegetation. The spores which the boiling has 
not been able to attack become transformed into 
adult bacteria which the heating of the next day will 
destroy. 

Heat acts with more rapidity upon bacteria in the 
moist condition than when dried, upon those of water 
than upon those of the air. 

Whether we have to do with adult germs or spores, 
with a dry medium or a moist, the duration of the 
heating is of great importance and can be supple- 
mentary to insufficiency of temperature. 

Antiseptics. This name has been given to chemical 
substances capable of destroying pathogenic germs or 



Transformation, etc., of Pathogenic Microbes. 101 

of arresting their development. The number of the 
antiseptics is quite considerable, but there are some 
which stand at' the top of the list and merit more es- 
pecially the confidence of the practitioner. These are, 
indeed, violent poisons for nearly all microbes, whilst 
the others, less energetic, are only really efficacious 
when they are directed against microbes the vitality 
of which is easily destroyed. Nevertheless, we can 
not establish a general rule upon this subject; the 
antiseptic action depends not only on the chemical 
agent employed, but also on the microbe concerned ; 
it is also dependent upon the duration of contact, the 
nature of the excipient, etc. The alcoholic solutions 
of carbolic acid, for example, are less antiseptic than 
aqueous solutions of the same concentration because 
the former have a smaller power of penetration than 
the latter. 

Antiseptic substances act either by rendering the 
media unsuitable for the multiplication of the mi- 
crobes, or by opposing themselves to the production 
of the diastases indispensable to the elaboration of their 
foods. 

We believe that it may be profitable to reproduce 
here the classification of the antiseptics proposed by 
M. Miquel. The antiseptic substances are here classi- 
fied according to the dose of the substances necessary 
for the sterilization of a litre of beef bouillon : 

1. The most powerful antiseptics : *01 to 0*1 gram. 
(Oxygenated water, corrosive sublimate, nitrate 

of silver.) 

2. Very strong antiseptics : 0*1 to 1 gram. 
(Iodine, bromine, sulphate of copper.) 



102 Manual of Veterinary Microbiology. 

3. Strong antiseptics: 1 to 5 grams. 
(Bichromate of potassium, chloroform, chloride 

of zinc, carbolic acid, permanganate of potas- 
sium, alum, tannin.) 

4. Moderate antiseptics : 5 to 20 grams. 
(Arsenious acid, boric acid, chloral hydrate, 

salicylate of sodium, sulphate of iron.) 

5. Weak antiseptics : 20 to 100 grams. 
(Borate of sodium, alcohol.) 

6. Very weak antiseptics : 100 to 300 grams. 
(Arseniate of potassium, iodide of potassium, 

sea salt, glycerin.) 



CHAPTER IV. 

METHODS OF DETERMINATION OF PATHOGENIC MICROBES. 

1. Examination, investigation, and staining of bacteria. 2. Cult- 
ure of bacteria. 3. Experimental inoculations. 

We have now, in order to finish the general study 
of pathogenic germs, to review the different methods 
of determination of these germs, that is, the means 
which we have of distinguishing them from each other, 
or recognizing them. This means consists in, 1st, mi- 
croscopical examination of the bacteria either directly 
or after coloring them; 2d, their culture in artificial 
media; and, 3d, the test of the effects which they de- 
termine in animals. 

In practice, and for certain germs which are well 



Methods of Determination of Pathogenic Microbes. 103 

characterized, the first of these means is often suffi- 
cient; but in other cases the germs to be determined 
must be subjected to the test of the three reactions 
which we have just mentioned. Thus, for example, 
the bacillus of human tuberculosis and that of avian 
tuberculosis possess identical physical characters, re- 
act in the same way to coloring matters, and give 
cultures difficult to differentiate ; on the other hand, 
they have different effects on experimental animals. 
In default of this last test we might have concluded 
that the two germs were identical when they are 
really quite distinct in some of their physiological 
properties. 

I. Examination, investigation, and staining of bacteria. 

The first examination of a liquid, from a bacterio- 
logical point of view, should always be made without 
the help of reagents ; we thus see the germs in their 
real form and size. Desiccation, or the addition of 
staining solutions often change the morphological 
character of microbes. It is in fresh preparations, 
also, that we are in a position to judge of the mobil- 
ity or immobility of the beings which we wish to 
study. 

Microbes being naturally colorless, little refringent, 
and consequently difficult to distinguish, staining 
methods have been adopted in order to bring them 
into relief in the media to be examined. 

For this purpose we have recourse to the colors de- 
rived from coal aniline colors which have an in- 
tense staining power. Without entering further into 
the details of this subject, we may say that, with the 
stains employed, we often associate adjuvant sub- 



104 Manual of Veterinary Microbiology. 

stances which -act the part of mordant, that is, which 
increase the penetrating power of the coloring sub- 
stance and render its fixation more energetic; such 
are, for example, alum, potash, soda, lithia, carbonate 
of ammonia, aniline, etc., all of which we will find 
in the various liquids used in histology and micro- 
biology. 

Heat is calculated to expedite the staining of mi- 
crobes, as well as of organic tissues in general; we 
shall see that this property has been taken advantage 
of in order to obtain rapid staining of various bacilli, 
and especially those of tuberculosis. 

Two groups of aniline colors are recognized. In 
the first (basic colors) the coloring substance is asso- 
ciated with a colorless acid. In the second (acid 
colors) it plays the part of acid and is combined with 
a colorless base. 

This distinction should be noted because the colors 
of the first class have a special affinity for cell nuclei 
and for bacteria, whilst those of the second give dif- 
fuse staining, that is, fix themselves indifferently on 
all parts of the cells and on the intercellular sub- 
stance. 

The basic colors include gentian and methyl violet, 
methyl green, methylene blue, fuchsin, bismark brown, 
vesuvin, and safranin. 

The principal acid colors are eosin and fluorescine. 

Single and double staining. In any preparation con- 
taining microbes we can give to all the elements, mi- 
crobic and non-microbic, the same coloration. In the 
case of certain micro-organisms we ought, indeed, to 
limit ourselves to this method of staining, such as 
the germs of fowl cholera, pneumo-eiiteritis of the 



Methods of Determination of Pathogenic Microbes. 105 

pig, etc. With these germs we can, at most, only 
attenuate the color of the tissue elements with the 
aim of bringing more distinctly into view the deeper 
color of the bacteria. Such is the principle of a pro- 
cess of single staining which has been applied with 
slight variations by different micro-biologists, and 
which will be further referred to later. 

Patient researches have placed at our command 
another method, called double staining. This method 
furnishes excellent preparations arid renders great 
service in the study of the distribution of microbes in 
the tissues. Double coloration is obtained by differ- 
ent means, of which the choice is not indifferent, for 
the same microbe does not take the stain by all of 
these processes. The manner in which microbes, be- 
have toward the methods of double staining is even 
utilized for characterizing them. 

The principle of double staining is as follows : 
1st, the whole preparation is stained uniformly; 2d, 
all the elements other than bacteria are decolorized 
whilst the latter retain, on the contrary, the color 
which was originally imparted to them ; 3d, the 
parts decolorized by the preceding operation are 
stained with a color which stands out distinctly from 
that of the microbe. Thus, the latter being violet, 
the contrast color will be red or brown; if, on the 
contrary, the microbes are stained red, the back- 
ground of the preparation should be stained blue or 
green. 

We may have to examine liquids, or solid tissues. 
In this last case it is sometimes necessary to make a 
previous examination of the organic pulp and study 
it later in thin sections. We shall therefore consider, 



106 Manual of Veterinary Microbiology. 

in succession, the various manipulations to which the 
liquids, pulp, and sections of organs must be sub- 
jected. 

Liquids Direct examination. A drop of the liquid 
is deposited upon a slide, covered with a cover glass, 
and at the margin of the latter is placed a drop of 
coloring matter ; penetration gradually takes place, a 
process which may be assisted by manipulation of 
the cover glass. "We may also add a trace of coloring 
matter directly to the drop of liquid to be examined 
before inclosing it under the cover glass. For this 
purpose aqueous solutions of gentian violet or methyl 
violet are generally employed. 

Examination after desiccation. The preparation just 
described has its field of usefulness, but it has the de- 
fect of not being permanent, and, especially, of not 
permitting of double staining. To attain these ends 
the liquids are rapidly dried so as to fix the elements 
which they contain and cause them to adhere to the 
glass. In this .process a drop of liquid is spread on 
a cover glass either by means of a spatula or by 
pressing the drop between two cover glasses which 
are glided upon each other and then separated. The 
cover glasses, thus coated, are held over the flame of 
a spirit lamp or placed upon a hot plate, the surface 
containing the liquid to be examined being directed 
upward so as not to be attacked by the flame. The 
drying is sufficient when, the liquid is transformed 
into an opalescent layer ; this layer then intimately 
adheres to the cover glass, which can be then trans- 
ported through a series of reagents without risk of 
the former becoming detached. 

The quantity of liquid employed should be very 



Methods of Determination of Pathogenic Microbes. 107 

small in order that the dried preparations may form 
a very thin coating, which, after staining, will not 
obstruct the passage of the rays of light. This 
recommendation is of special importance in the case 
of organic fluids containing many histological ele- 
ments, such as pus, for example, of which only a very 
small particle should be taken. 

Before all staining, it is sometimes necessary to re- 
move the fatty matters from the dried substance by 
immersing it in chloroform or a mixture of equal 
parts of alcohol and ether; this practice is indispensa- 
ble in operating upon milk. 

Preparations of dried blood are also much im- 
proved by this treatment : under the influence of the 
alcohol and ether mixture the corpuscles acquire a 
stability which desiccation can not confer upon them, 
and they then present very distinct forms in stained 
preparations, in which their relations with the mi- 
crobes can be studied. This is a highly commenda- 
ble practice which much increases the precision of 
researches bearing upon the blood. It should be fol- 
lowed by a second drying. 

Organic pulps. These are obtained by scraping the 
freshly cut surface of the organ suspected of contain- 
ing microbes ; a small particle is spread out between 
two cover glasses in a thin layer, in the manner indi- 
cated above, and then dried. 

Microscopic sections. Tissues which are intended to 
be cut for the study of the distribution of the mi- 
crobes which may be present, should, first of all, be 
well fixed, that is, their elements well immobilized as 
regards their form and relations, and thus placed be- 
yond the reach of cadaveric changes. This is es- 



108 Manual of Veterinary Microbiology. 

pecially necessary in the examination of those tissues 
for micro-organisms, as these latter, living independ- 
ently, may continue to multiply after the death of 
the tissues, and, further, in organs left in contact 
with the air foreign germs generally develop which 
are prejudicial to the accuracy of such researches. 
These accidents are avoided hy immersing the tissues 
to be preserved in absolute or, at least, strong alco- 
hol, the precaution having been taken, in order to 
render the action of the alcohol more rapid and more 
intimate, to cut the organs into small cubes of half 
to one centimeter on the side. 

Sectioning is performed in various ways, but the 
technique of this operation can not be described 
here ; its study belongs to the domain of histology. 
Moreover, this mode of research does not appear to 
be within reach of all practitioners on account of the 
instruments and time which it requires, and, in 
nearly all cases, it can be omitted when the differen- 
tial diagnosis of infectious diseases is the only object 
in view. 

Mounting of preparations. 

Cover glasses. After staining, the cover glasses 
must be dehydrated to admit of their being mounted 
in Canada balsam. This end is attained very rapidly 
by drying them afresh over the flame of the spirit 
lamp or upon the hot plate, after having previously 
drained them between two folds of filter paper. As 
soon as the cover glass is dried, the preparation is 
cleared by depositing successively upon the coated 
surface a drop of cedar oil, clove oil, or bergamot oil, 
a drop of xylol, and, finally, a drop of Canada bal- 
sam dissolved in xylol. The cover glass is then 



Methods of Determination of Pathogenic Microbes. 109 

placed upon the slide in such a manner that the bal- 
sam distributes itself between the two glasses. 

We can omit having recourse to clearing (clove oil, 
cedar oil, etc.), and, after drying, pass directly to the 
balsam ; but it is incontestable that, if this practice 
increases a little the duration of the work, it also 
gives more distinctness to the preparation. 

Sections. Stained sections must also be dehydrated 
in order to be mounted in balsam. For this purpose 
they are passed successively through specimens of 
alcohol of different strength, ending with absolute al- 
cohol (70, 90, 100 per cent). They are then cleared 
by immersing them in one of the substances men- 
tioned above (cedar oil, bergamot oil, clove oil), then 
placed in xylol, and, finally, on a drop of balsam 
which has previously been deposited on a slide and 
which is now at once covered by the cover glass. 

These diverse operations may be made upon the 
slide itself, the stained section having been spread 
and fixed at the start by a semi-desiccation. This 
last is easily accomplished either by the aid of filter 
paper which is used to absorb the water from the 
section, or by moderate heating. 

Methods of single staining. 

. Hydro-alcoholic solutions of aniline colors. These are 
.made extemporaneously by adding a few drops of 
a saturated alcoholic solution of the dye to a watch 
glass of distilled water. They are largely used for 
single staining, especially for cover glasses. This 
method is sufficient for a rapid examination, when 
the object is merely to determine the presence or ab- 
sence of microbes in the preparation. 



110 Manual of Veterinary Microbiology. 

The solutions most used are those of gentian violet, 
methyl violet, fuchsin, and methylene blue. A few 
minutes contact is sufficient : five for cover glasses, 
fifteen for sections, a little longer in the case of 
methylene blue. 

Loffler's method. Cover glasses are stained for from 
five to ten minutes in the following medium : 

Solution of caustic potash, 1 to 10,000, 3 cub. cent. 

Saturated alcoholic solution of methyl- 
ene blue, . . . . 1 " " 

All the elements are stained deep blue. To bring 
out the microbes, the stained preparations are im- 
mersed in water containing a small quantity of acetic 
acid, one or two drops to a watch glass ; in this they 
remain on an average only one minute; they are then 
washed with distilled water and mounted. The mi- 
crobes appear colored a deep blue, the tissue ele- 
ments a light blue. 

Method of Malassez and Vignal. Staining is here 
made with Malassez' blue, which consists of: 

Aniline water, .... 9 cub. cent. 

Absolute alcohol, . . . . 1 " " 

Saturated alcoholic solution of methyl- 
ene blue, . . . . . 1 " 

Sections remain therein ten minutes, cover glasses 
five minutes ; they are then decolorized in the follow- 
ing mixture: 

Absolute alcohol, .... 1 cub. cent. 

2 per cent solution of sodium carbonate, 2 " " 

Methods of double staining. 
Gram's method. Gram's violet contains: 
Aniline water, . . 10 cub. cent. 



Methods of Determination of Pathogenic Microbes. Ill 

Absolute alcohol, . . . .1 cub. cent. 

Saturated alcoholic solution of gentian 
violet, 1 " " 

Cover glasses remain in this fluid five minutes, sec- 
tions fifteen, after which they are transferred for one 
to two minutes into G-ram's solution of iodine in iodide 
of potassium which consists of: 

Iodine, 1 gram. 

Iodide of potassium, ... 2 grams. 

Distilled water, .... 300 grams. 

They are then completely decolorized in absolute 
alcohol, and the ground- work of the preparation 
stained red (eosin, picro-carmine) or brown (bismark 
brown, etc.) The violet remains fixed on the mi- 
crobes, but in the tissue elements it is replaced by 
the red or brown. 

Weigerfs method. Weigerfs violet consists of : 

Saturated aqueous solution of methyl violet 

6 B, . . . . . .68 grams. 

Absolute alcohol, . . . . 11 grams. 

Aniline, .3 grams. 

Sections are stained from five to ten minutes, im- 
mersed for one to two minutes in Gram's iodine 
solution, passed rapidly through absolute alcohol (a 
few seconds) in order to remove the greater part of 
the water, and decolorized in aniline oil; they are 
then cleared in xylol and mounted in balsam. Since 
by this method contact with alcohol is avoided, it is 
necessary to stain the groundwork of the preparation 
in red or brown before staining with violet. This 
inversion may also be employed without inconven- 
ience in staining by the Gram method. The employ- 
ment of alcohol in dehydrating may be entirely 



112 Manual of Veterinary Microbiology. 

avoided in another way; to this end, the section, after 
coming from the iodine, or even before staining, is 
spread out, moist, upon the glass and fixed there in 
a state of semi-desiccation by slightly heating it or 
by the aid of tissue paper. On the section, thus 
fixed, are then deposited in succession a drop of the 
different reagents, the excess of each being carefully 
removed with the tissue paper before replacing it 
with the next. 

In the methods of Gram and Weigert as well as in 
the method by Kiihne's violet, which we will consider 
later, the iodine solution of Gram may be replaced by 
the following : 

Corrosive sublimate, ... 1 gram. 

Water, . . . . . .100 grams. 

Alcohol, enough to dissolve. 

All microbes do not take indifferently the colora- 
tions obtained by the above methods. For a given 
germ one method may be more'suitable than another; 
the manner in which the microbes behave toward the 
different methods of staining is, indeed, utilized in 
distinguishing them. But it is often desirable first 
of all to determine whether or not a given substance 
contains any germs. We attain this end by the fol- 
lowing method : 

Kuhne's method. Kiihne has succeeded in stain- 
ing nearly all microbes by employing two processes 
in succession, one with methylene blue, and the other 
with crystal violet. Some microbes are colored only 
by the blue (typhoid bacillus), others only by the 
violet (bacillus tuberculosis, bacillus of leprosy) ; 
others, again, and these are the most numerous, take 
the two colors indifferently. 



Methods of Determination of Pathogenic Microbes. 113 

This method is therefore useful for determining the 
presence of microbes in any tissue. In its execution 
an equal number of sections are treated by both 
methods. 

1. Method by Ratine's blue. The blue of Kilhne is 
obtained by adding, drop by drop, to a one per cent 
solution of carbonate of ammonia, a saturated aque- 
ous solution of methylene blue until the mixture ac- 
quires a deep blue color. The sections, after dehy- 
dration in alcohol, remain ten minutes in this blue 
liquid, are then rapidly decolorized (two or three sec- 
onds) in a 1 to 1000 aqueous solution of hydrochloric 
acid, then passed into distilled water to remove all 
trace of the acid. They are afterward mounted with- 
out passing through alcohol, which would remove 
too much of the stain. For this end, after removal 
from the water, they are spread out on a slide and 
there allowed to dry, or better, the desiccation is has- 
tened either by means of a current of air or very 
slight heating. They are then cleared in xylol and 
mounted in balsam. 

2. Method by crystal violet. Kuhne's violet is obtained 
in the same way as the blue solution by adding sev- 
eral drops of a saturated aqueous solution of crystal 
violet to a one per cent aqueous solution of carbonate 
of ammonia until a deep violet color is produced. 
Sections, after dehydration in alcohol, are left in this 
solution ten minutes (two hours for the bacillus tuber- 
culosis), washed in distilled water, and immersed for 
two to three minutes in Gram's iodine solution ; from 
this they are transferred into a saturated alcoholic 
solution of fluorescine until decoloration is nearly 
complete. The remaining coloring matter is extracted 



114 Manual of Veterinary Microbiology. 

in absolute alcohol, the sections are cleared in oil of 
cloves, passed into xylol, and finally mounted in bal- 
sam. 

To obtain double staining by the violet method it 
suffices to color the sections with picro-carmine be- 
fore placing them in the violet solution. 

Kiihne has modified this process so as to avoid the 
employment of alcohol and oil of cloves. The sec- 
tions, after dehydration in alcohol, are immersed for 
ten minutes in the concentrated aqueous solution of 
crystal violet, to which has been added hydrochloric 
acid (one drop to fifty grams of the solution). They 
are washed in distilled water, treated with Gram's 
iodine solution, replaced in water, then passed rapidly 
(a few seconds) through absolute alcohol and into 
aniline oil in which they are decolorized. The ani- 
line is removed by xylol, and the sections mounted in 
balsam. The employment of alcohol can be entirely 
avoided by transferring the sections after the action 
of the iodine on to the slide and there treating them, 
after dehydration, with aniline, xylol, and balsam, in 
succession. 

Method of Berlioz. This gives a rapid double color- 
ation. The sections are left for a quarter of an hour 
in a mixture of equal parts of the two following 
solutions: 

1. Distilled water, . . . / 84 cub. cent. 
Aniline water, .... 6 " 

Methyl violet 6 B, ,. , . , . 2-5 grams. 
Absolute alcohol, . . . .10 cub. cent. 

2. Distilled water, . ... 95 " 
90 per cent alcohol, 5 " 
Coccinine, - 2-5 grams. 



Methods of Determination of Pathogenic Microbes. 115 

They are then passed through the Gram iodine 
solution, or a five per cent solution of sodium car- 
bonate, through water, alcohol, etc. This method is 
well adapted for the staining of charbon material. 

Method of staining spores in bacilli. To stain spores 
dried on the cover glass they are immersed fol 1 a few 
minutes in hot Ehrlich's solution and washed in 
alcohol; the cover glass is then held for a second 
in methylene blue, washed again in alcohol, and 
mounted in balsam. The bacilli are blue, the spores 
are red when they have attained their development, 
uricolored whilst they are in way of formation. 

II. Culture of germs. 

The majority of pathogenic germs can be culti- 
vated in appropriate artificial media placed under 
suitable conditions of temperature. Our aim'not be- 
ing to publish a complete treatise on the technique of 
bacteriology, we will refer the reader for details upon 
this point to special works on the subject, and limit 
ourselves to the general principles of the question. 

We will review, in succession, the methods of ster- 
ilization, the composition and preparation of the va- 
rious culture media, the sowing of these media and 
establishing them at a suitable temperature, the physi- 
cal and chemical characters which the cultures may 
present and the preservation of these cultures. 

Sterilization. 

The objects and instruments which are employed 
in the culture of microbes must be previously freed 
from all living germs. Sterilization is obtained by 
different means : heat, filtration, antiseptics. 



116 Manual of Veterinary Microbiology. 

Heat. As we already know, bacteria and their 
spores are killed by heat. For this it is necessary 
that the objects which contain the germs be raised to 
a temperature varying according to the case. There 
are several processes. 

a) Flaming. Objects which -can support the ac- 
tion of a rather high temperature are brought into 
the flame of a spirit lamp and subjected, in all their 
parts, to a temperature of about 200. Flaming is 
practiced chiefly for glassware, occasionally for metal- 
lic instruments : spatulas, platinum wire. 

b) Dry air ovens at 150 C. The objects to be steril- 
ized are kept for about two hours at a temperature 
of 150. To this end they are placed in appropriate 
ovens, furnished with a temperature regulator and 
arranged in such a manner that the temperature, 
through the circulation of hot air, is nearly the same 
in all parts of the apparatus. These ovens are pro- 
tected from too free radiation by walls composed of a 
substance which is a bad conductor of heat, such as 
asbestos. Sterilization by this process is practiced 
chiefly on glassware, wadding and metallic objects. 

c) Koch's steam sterilizer. This is a generator of 
steam, of cylindrical form, containing water which is 
kept in ebullition by the combustion of gas ; it is sur- 
mounted by a dome of plate tin enveloped in felt to 
prevent cooling. The temperature of the cylinder, 
charged with steam, is maintained at 100, a tempera- 
ture sufficient for the destruction of germs and spores 
in the moist state. Objects to be sterilized must be 
left in the upper part of the apparatus from two to 
three hours. The steam sterilizer is employed chiefly 



Methods of Determination of Pathogenic Microbes. 117 

for the sterilization of culture media, and for objects 
which can not sustain a temperature of 150. 

d) Papin' s steamer The Chamberland autoclave. 
Sterilization of objects in fluid media is much more 
rapid and certain when they are subjected during ten 
to fifteen minutes to a temperature of 120. To 
realize these conditions the objects are inclosed in a 
Papin steamer, which admits of obtaining, under 
pressure, the temperature indicated. 

Chamberland has recently devised an autoclave or 
steamer specially adapted for sterilization. It is now 
found in all laboratories. It is merely a Papin 
steamer modified for this particular use. 

e) Discontinuous heating. Some media, such as 
blood serum and milk, can not endure a temperature 
of 100 without undergoing considerable modifica- 
tions. For the sterilization of these substances we 
have recourse to the method of Tyndal, which con- 
sists in killing the germs in full vegetation, an ope- 
ration which only requires a temperature of 58, 
maintained during two hours. For blood serum the 
temperature should not exceed 58, for this medium be- 
comes coagulated at a higher temperature. The first 
heating not having killed the spores, the substance is 
then brought to the ordinary room temperature, or 
better, to 37; the spores then quickly vegetate and 
the bacteria .to which they give birth will be killed 
by a second heating on the following day. This ope- 
ration, repeated four or five times, ends in the certain 
sterilization of the liquid. 

The apparatus employed in this method of steril- 
ization consists of water baths maintained at a con- 
stant temperature by a regulator. 



118 Manual of Veterinary Microbiology. 

Filtration. Germs, like all solid particles, can not 
pass through porous substances in wh'ich the pores 
are very fine. Liquids filtered through such a sub- 
stance are therefore sterilized. Hence, it is possible 
to obtain sterilization by filtering through plaster of 
Paris, amianth, or porcelain. 

Chamberland has constructed a filter for steriliza- 
tion based upon this property. It consists of a hol- 
low tube of porcelain, closed at one end; this tube is 
immersed in the liquid to be sterilized and a vacuum 
produced in its interior in any of the ordinary ways ; 
the sterile liquid passes into the interior of the tube. 
The latter, of course, should have been previously 
sterilized and be free from fissures. 

Antiseptic solutions. Sterilization of instruments, 
anatomical specimens, the hands, etc., should be ob- 
tained by means of acidulated solutions of corrosive 
sublimate at 2 to 1,000, carbolic acid at 5 per cent, or 
creolin at 2 per cent. To obtain sterilization, the 
objects, after thorough cleansing, must be washed 
with these solutions. 

Culture media. 

The culture of germs in artificial media necessitates 
the presence in these media of all the principles es- 
sential to their life, as well as the absence of all 
noxious products. Each germ having special nutri- 
tive requirements of its own, the ideal would be to 
possess media especially appropriate for the diverse 
pathogenic species. A perfect culture apparatus 
ought to admit of the continual addition of nutri- 
ment, with, at the same time, the elimination of the 
residual products ; but this ideal is far from being 



Methods of Determination of Pathogenic Microbes. 119 

realized : the nutritive media are, for the most part, 
prepared empirically, starting from complex animal 
or vegetable products, and not only are we unac- 
quainted with the qualitative and quantitative com- 
position of these media, but we are also ignorant of 
the special nutritive value of the essential principles 
they contain. Moreover, our culture apparatus are 
very imperfect; during the progress of the culture 
the nutritive material becomes exhausted and charged 
with the products of nutrition, products which are 
prejudicial to the normal life of the germs. From 
theee circumstances it results that artificial cultures 
do not always give individuals conforming with 
those which are sowed, and that often, in the case of 
pathogenic species, they furnish degenerated beings 
incapable of acting upon man or animals. 

Generally speaking, all culture media ought to con- 
tain, besides the necessary nutritive substances, at 
least sixty per cent of water, they should be neutral 
or slightly alkaline, and absolutely sterile. A con- 
siderable number of media are in use; they embrace 
organic liquids : milk, urine, blood serum ; various 
decoctions : bouillons of meat, hay, fruits, beer wort, 
etc. These materials are used alone or after the ad- 
dition of supplementary nutritive substances : pep- 
tone, gelatin, glycerin, glucose, phosphates, etc. 

Without entering into the details of the prepara- 
tion of these diverse media we will describe those 
most commonly in use. 



120 Manual of Veterinary Microbiology. 

A. FLUID MEDIA. 
1. Bouillons. 

Bouillons are the media most commonly used.* 
They are prepared from the flesh of the different do- 
mesticated animals, but especially from that of the 
calf, the ox, and the chicken. 

One kilogram of lean meat, free from bone, is 
finely minced and allowed to macerate for twenty-four 
hours in two liters of water, in a cool place. The 
reddish fluid which bathes the meat is expressed, 
brought up to its original volume, and to it is added 
one-half per cent of table salt and a trace of potas- 
sium phosphate, occasionally, also, from one to three 
per cent of peptone, glycerin, and glucose. The 
liquid is then cooked for an hour at 100 in the Koch 
sterilizer, by which means a certain amount of its al- 
bumen is coagulated; it is strained through linen and 
then neutralized with a one per cent solution of 
caustic potash, or a five per cent solution of sodium 
carbonate. The filtered and neutralized bouillon is 
subjected, during ten minutes, to 115 in the auto- 
clave, then again filtered. This "bouillon, introduced 
into a conical flask stopped by a plug of wadding, is 
finally sterilized by a last heating of a quarter of an 
hour at 115. It is then ready to be introduced into 
the culture vessels. The culture vessels most com- 
monly employed are the Pasteur bulbs. These are 
small vials of thin glass, flat on the bottom, and with 

* [Except for special purposes, or where a large quantity of the 
culture is required, solid media are now more in use than bouil- 
lons. D.] 



Methods of Determination of Pathogenic Microbes. 121 

a very short neck, which is surmounted by a ground 
glass hood ; the latter is prolonged by a narrow tube 
which is closed by a plug of wadding. These ves- 
sels may be replaced by conical vials, etc., but the 
latter have the great inconvenience of allowing a too 
rapid evaporation of the liquid. 

In filling the Pasteur vessels, which must previously 
have been sterilized, we employ with advantage a 
Chamberland pipette. This is a flat bottomed bulb 
\vith a long narrow tube bent to 45 and filled at 
one part of its length by a plug of wadding; from 
the side of this bulb projects horizontally a long slen- 
der tube sealed with the flame at its free extremity. 
The pipette having been sterilized, the lateral tube 
is flamed, the point broken off in the sterile bouillon 
and the latter slowly aspirated into the pipette. This 
done, a quantity (half to one centimeter in depth) of 
the bouillon is allowed to flow into the Pasteur 
culture vessels, the latter being held in the horizontal 
position, in order to avoid the entrance of air germs; 
this operation should be conducted in a room free from 
currents of air. The bouillon having been introduced 
into the culture vessels, it is advisable to test if these 
are really sterile by placing them during two or three 
days in an incubator ; those in which the bouillon be- 
comes turbid contain bacteria and consequently must 
be rejected.* 

* [Usually the culture media, both fluid and solid, are filled into 
ordinary test-tubes, closed with cotton and sterilized in the steam 
sterilizer. Evaporation is to a large extent prevented by accu- 
rately fitting a piece of tin-foil over the mouth of the tube. D.] 



122 Manual of Veterinary Microbiology. 

2. Milk. 

Milk may be collected in a state of purity in ster- 
ilized tubes by introducing a sterile canula into the 
teat after the latter has been well disinfected. This 
is the best means of obtaining milk free from germs, 
but it is one which is not generally available, and, 
most frequently, we are compelled to sterilize this 
liquid. 

This is done in the autoclave at 115. The milk, 
without the cream, is then introduced into the Pas- 
teur bulbs and subjected to the test of the incubator. 

3. Urine. 

It was in urine that Pasteur first cultivated the 
bacteridium of charbon. The urine, after collection, 
is rendered alkaline, filtered, and sterilized exactly 
like the bouillon. It may be employed alone or after 
the addition of supplementary nutritive substances. 

B. SOLID MEDIA. 

This consists of fluid media gelatinized, coagulated 
blood serum, potatoes, etc. 

Gelatinized bouillons. 

Gelatinizing substances are added to bouillon, sim- 
ple or complex, so as to render it solid and trans- 
parent on cooling. The substances employed are 
gelatin, gelose, iceland moss, etc. 

Nutritive gelatin. To the bouillon whilst in prepa- 
ration and before boiling is added ten per cent of 
gelatin. The fluid is passed through a cloth, neutral- 
ized, and the process generally conducted as described 



Methods of Determination of Pathogenic Microbes. 123 

for bouillon. The product should be of a citron color 
and perfectly transparent after filtration. It is then 
introduced into test tubes plugged with wadding, and 
sterilized. These tubes, filled to one-third of their 
height, are then placed in a wire cage and kept in 
the autoclave at 105, during ten minutes. The 
gelatin solidifies on cooling and the tubes are then 
ready for use. 

Gelose or agar-agar. This is a gelatinous substance 
coming from certain algse of the Indian Archipelago. 

It is added to the bouillon whilst in preparation, in 
the proportion of one to two per cent, and the further 
process conducted as has already been described for 
gelatin. The addition of the agar often renders the 
liquid turbid, and this turbidity persists in spite of 
filtration; it is with the view of obviating this in- 
convenience that it has been recommended to inti- 
mately mix with the mass, after cooling to 50, the 
white of an egg beaten up, and then bring the whole 
to ebullition again. In coagulating, the albumen 
carries with it all the substances in suspension and 
the product becomes clearer. However it always re- 
mains slightly opalescent. 

The filtration of this liquid, like that of gelatinized 
liquids in general, ought to be performed while hot, 
and it requires some time. A hot filtering apparatus 
is employed, consisting of a glass funnel contained in 
a larger one of copper and separated from this last 
by a space full of water kept at the temperature at 
which the gelatinous mixture becomes fluid. The 
filtration thus obtained is slow, for the gelatin dries 
and hardens upon the filter where it contacts the 
walls of the glass funnel. 



124 Manual of Veterinary Microbiology. 

For filtrations of this kind we have employed with 
advantage a special support bearing a simple glass 
funnel provided with a lid and adjusted above a con- 
ical vessel. The gelatinous mass .to be filtered hav- 
ing been introduced into the funnel upon an ordinary 
filter, the whole is placed in the Koch steam sterilizer. 
We thus obtain a rapid filtration without desiccation 
or loss of the fluid, since the filtration takes place in 
an atmosphere of steam. The agar and gelatin hav- 
ing been poured into tubes and sterilized, the latter 
are placed in the cold, some in vertical position, others 
inclined so as to distribute the mass in a very large 
oblique layer; either the surface of this layer or the 
depth, in the vertical tubes, serves as a field for cul- 
ture. 

Agar has several advantages over gelatin : it is not 
fluidified by the growth of germs, it remains solid at 
40, and, therefore, admits of cultures in the incu- 
bator at 39; it may be subjected to cooking for a 
long time without losing its gelatinizing power. On 
the other hand, it has the disadvantage of being al- 
ways slightly cloudy, is not well adapted for plate 
cultures, and gives cultures which are not well de- 
fined. 

Agar-gelatin. With the aim of making plate cul- 
tures at the temperature of 39, Jensen has recom- 
mended, for the solidification of the bouillon, a mix- 
ture of agar and gelatin. He adds to the bouillon 5 
per cent of gelatin and 0.75 per cent of agar. The 
preparation is very clear, it is liquefiable at a tempera- 
ture which does not kill the germs, and it remains 
solid at 39. 



Methods of Determination of Pathogenic Microbes. 125 

Gelatinized serum. 

A solid medium of frequent use is sterilized and 
cooked blood serum. Many methods are employed 
. in collecting and sterilizing the serum ; these we re- 
frain from describing here and limit ourselves to a 
description of that which we are accustomed to use. 
Blood drawn from the jugular of the horse or the ox 
is collected in deep cylindrical glass vessels and left 
in the cold for twenty-four hours. The clot contracts 
and the serum comes to the surface; the latter is 
collected by means of a pipette and introduced into 
conical vials to be sterilized. 

Sterilization is obtained by conveying the serum 
on several successive days into a water bath a 58, 
and, each day, leaving it there for two hours. The 
bath is provided with a thermo-regulator by which a 
temperature of 58 is quickly obtained, a condition 
indispensable to prevent the germs from multiplying 
too much and, in consequence, altering the coagula- 
bility of the serum. After eight days heating the 
liquid maybe considered sterile ; it is then distributed 
in test tubes by means of a Chamberland pipette. 
Other nutritive substances, peptones, glycerin, etc., 
may be added to the serum before sterilization. 

The cooking of the serum takes place in double 
walled ovens so arranged that the tubes may be in- 
clined so as to spread out the liquid. These ovens 
are regulated to 70; the tubes are removed as the 
serum in them becomes coagulated. Before being 
used these tubes are tested during three days in the 
incubator. 



126 Manual of Veterinary Microbiology. 

Potatoes. 

Some germs admit of cultivation upon the potato. 
For the preparation of this medium sound tubers 
with smooth skin are selected, thoroughly washed in 
water with a brush, and allowed to soak some time 
in 1 to 1,000 sublimate solution. They are then 
cooked by steam at 100, or in the Chamberland auto- 
clave, cut in two with a previously flamed table- 
knife, and deposited in a moist chamber. 

A moist chamber is usually represented by two 
crystallizers of different sizes, and well disinfected; a 
piece of filter paper saturated with sublimate solution 
is placed in the smaller, which is then covered by the 
larger. We thus obtain a moist space protected from 
the atmospheric germs. 

Sometimes potatoes, cut in sections, are cooked in 
tubes ready for culture ; these have a constriction 
near the base to retain the potato about the middle 
of the tube. 

Methods of culture. 

In order to cultivate germs we must first possess 
pure sowings. Occasionally the germ may be ob- 
tained pure from the organism in which it has de- 
veloped, as, for example, in bacteridian charbon, 
chicken cholera, rouget of the pig, etc. 

In most cases the micro-organisms do not occur in 
a state of complete purity in the parts of the organ- 
ism in which they pullulate ; it then becomes neces- 
sary, first of all, to separate them from other germs. 
We have, therefore, to describe, successively, the 
methods of isolation of germs, the inoculation of the 



Methods of Determination of Pathogenic Microbes. 127 

various media, and the establishment of a suitable 
temperature. 

Methods of isolation. 

The isolation of mixed germs may be obtained in 
several different ways. One method of isolation is 
based upon the different properties of the species to 
be isolated. Thus, the germs of the septicaemias of 
the rabbit and of the mouse have been withdrawn 
from the mass of microbes which pullulate in blood 
in way of putrefaction, by taking advantage of the 
fact that these germs are pathogenic for such 
animals. 

Pasteur has isolated the bacillus anthracis from 
the septic vibrio in charbonous blood in a state of 
putrefaction by taking into account the fact that the 
former is aerobic and the latter anaerobic : cultures 
in the air give the bacillus anthracis only; those pro- 
tected from oxygen give the septic vibrio. 

The influence of various chemical agents and that 
of heat at different degrees are also effectual means 
of isolation ; they kill some germs or considerably in- 
terfere with their multiplication, and thus favor the 
predominance of other less sensitive germs. 

Klebs has succeeded in isolating germs by basing 
himself upon their unequal distribution in the liquids 
in which they pullulate. Some, immobile, are found 
at the bottom or on the walls of the vessels ; others, 
motile, are uniformly distributed; some again are 
very greedy of oxygen and are found in the super- 
ficial layer. By withdrawing germs from these vari- 
ous parts more of one species than of another will 
be removed, and by repeating the operation several 



128 Manual of Veterinary Microbiology. 

times in succession one species will be obtained in a 
pure state. 

Cohn has separated certain germs by taking advan- 
tage of the resistance of their spores to a few mo- 
ments ebullition. The bacillus subtilis, or hay bacil- 
lus, is obtained by boiling neutral infusion of hay and 
then transferring to the incubator. The spores of 
the bacillus subtilis alone develop because these alone 
have resisted. 

Germs can be isolated by more direct means. 

Method of isolation by Salmonseri's capillary tube. 
This author has studied, in this way, the various 
germs which grow in putrefactive blood. He aspirated 
defibrinated blood into long capillary tubes which he 
closed at both ends and fixed upon a horizontal card. 
The few germs which are included in the tube grow 
separately in the different parts of its length ; their 
evolution can thus be studied and they can be col- 
lected separately for cultivation. 

Method of isolation by dilution. A material, rich in 
germs of different kinds, is diluted with sterilized 
water so that the germs are considerably rarefied. 
This liquid is then inoculated by drops into a series 
of culture vessels ; those of these vessels which have 
received only one germ will give a pure culture on 
incubation. 

MM. Roux and Yersin have succeeded ip isolating 
the bacillus of human diphtheria by making with a 
platinum wire charged with a trace of false mem- 
brane, longitudinal streaks in a series of tubes of 
blood serum. The germs become progressively rare- 
fied on the wire and ultimately the serum in the last 



Methods of Determination of Pathogenic Microbes. 129 

inoculated tubes receives no more than one germ; 
the culture is then pure. 

Method of isolation by plate cultures. Koch has re- 
commended a method of plate culture now practiced 
in all the laboratories. A trace of the substance 
containing the bacterial mixture is inoculated to a 
tube of gelatin which is then fluidified by heating at 
30. The gelatin is then agitated so as to uniformly 
distribute the germs throughout its substance, and 
poured upon a sterilized glass plate which is placed 
horizontally on a cold surface, and covered with a 
bell jar. The gelatin spreads out in a thin solid layer 
in which the germs find themselves isolated. The 
plate is then placed on a small bench in the moist 
chamber, in a room kept at a temperature of about 
20. Each germ gives an isolated colony, the appear- 
ance of which can be studied and from which seed 
can be obtained for starting new cultures. In this 
process the germs which grow on the surface of the 
layer of gelatin and which come, for the most part, 
from the exterior should not be taken into account. 

Instead of using a plate the gelatin may be spread 
out in the tube itself or in any vessel presenting a 
very large interior surface.* This mode of procedure 
has the advantage over the original technique that it 
exempts the culture from all risk of external infection 
and allows of anaerobic cultures. 

The substitution of agar-gelatin for gelatin consti- 

* [The original " plate process " of Koch is now, for the most 
part, supplanted by the more convenient and safer method of 
Petri, in which the fluidified gelatin, after inoculation in the test 
tube, is poured into a shallow glass dish provided with a cover 
(crystallizers) . D.] 



130 Manual of Veterinary Microbiology. 

tutes another improvement, the first of these sub- 
stances admitting of being raised to 39 without 
fluidifying. The temperature of 39 is much more 
favorable for the vegetation of germs than that of 20. 

Inoculation of culture media. 

Inoculation of culture media should be performed 
in such a manner as to avoid the introduction into 
the culture medium of external germs, either from 
the air or the surface of external objects. Air germs 
will be avoided by considerably inclining the bulb or 
tube to be inoculated toward the horizontal and ope- 
rating rapidly but quietly so as not to cause agitation 
of the air. The instruments used in transferring the 
seed must first of all have been disinfected. These 
instruments are : 

1st. A somewhat rigid platinum wire, three to five 
centimeters in length, fused to the end of a glass 
rod of small diameter. This wire is brought to a red 
heat in the flame of an alcohol lamp ; as soon as it is 
cooled the extremity is charged with a trace of the 
seed which is then transferred to the medium to be 
inoculated, shortening as much as possible the course 
to be traversed by the wire in order to diminish the 
chances of infection from the air. In the case of 
solid media the seed may be deposited in a line on 
the surface or in a vertical track into the substance 
of the gelatinized mass. The appearance of the cul- 
ture will naturally vary with these two modes. 

The platinum wire may be replaced by a needle of 
glass, readily obtained by drawing out a fusible glass 
rod over the lamp. 

2nd. Capillary tubes. A glass tube is drawn out in 



Methods of Determination of Pathogenic Microbes. 131 

the flame so as to reduce it to capillary dimensions; 
the capillary segment which has been perfectly ster- 
ilized by the heat is then closed at its two ends. 
When used, it is flamed, the two ends broken off, 
and one of them introduced into the liquid contain- 
ing the seed, which ascends by capillarity: the tube 
is then passed into the culture medium and a drop 
of the fluid expelled by blowing at the other end. 
Such a tube can only be used once. 

3d. Pasteur 's pipette. Pasteur has designed a special 
pipette for collecting and sowing, in a state of purity, 
liquids containing germs. It consists of a glass tube 
five or six centimeters in length and about one centi- 
meter in diameter; this tube is drawn out and closed 
at one of its extremities the other being provided 
with a constriction, and filled with wadding; the 
whole is sterilized at 150. In using it, the slender 
end is flamed, the point broken off, and the fluid con- 
taining the seed, blood, various serosities, urine, pus, 
cultures, etc. aspirated into the tube. The pipette is 
then introduced into the medium to be inoculated and 
one or two drops allowed to flow out. A liquid con- 
taining bacteria may be preserved in this tube for 
some time by taking the precaution to seal the capil- 
lary extremity in the flame. The Pasteur pipette can 
be used for sowing fluid media only ; the platinum 
wire serves equally well for both fluid and solid 
media. 

Placing at a ^suitable temperature. 

Whilst most germs are able to grow at the tem- 
perature of 15 to 20 C., many of them grow better 
at a temperature of 30 to 40, and a certain number 
are able to grow at this temperature only. 



132 . Manual of Veterinary Microbiology. 

For cultivation at 20 it is sufficient to place the 
inoculated media in an ordinary room ; this room 
should be heated in winter, and in summer, on the 
other hand, should be protected from a too free en- 
trance of the sun's rays. If necessary, a room can 
be provided with double windows and padded door, 
in which is kept a stove receiving gas from a thermo- 
regulator placed in the room ; a constant temperature 
can thus be obtained. 

"When the cultures require a temperature of from 
30 to 40, we have recourse to special ovens. These 
are air ovens in which a suitable temperature is main- 
tained. 

Culture ovens are of various forms and sizes. 
They may have single or double walls; in the latter 
case the space between these walls contains a layer 
of water. They are, further, furnished with an ar- 
rangement which permits of the renewal of the in- 
terior air. These ovens may be regulated for differ- 
ent temperatures; usually they are heated to 39 C. 
Culture ovens have also been constructed, the tem- 
perature of which varies at different levels but re- 
mains constant for each of these. 

Two principal conditions must be fulfilled in order 
to maintain a constant temperature ; the loss of heat 
must be reduced to a minimum and be invariable, 
and the heat communicated to the apparatus must be 
equal to the heat lost. The first condition is obtained 
by surrounding the ovens with a body which is a bad 
conductor of heat, such as felt, sometimes by a 
double wall and a double door, and by placing them 
in a room kept at a uniform temperature. As to the 
second, it requires a more complicated arrangement; 



Methods of Determination of Pathogenic Microbes. 133 

the gas taken from the pipe passes first into a regu- 
lator which corrects the variations of pressure at the 
gasometer at different periods of the day, and then 
arrives at the apparatus hy passing through a tem- 
perature regulator. This last is of various forms; it 
is usually a kind of mercurial thermometer sur- 
mounted hy a chamber into which the gas enters; 
when the temperature rises in the oven in which it 
is placed, the mercury, in expanding, partially ob- 
structs the entrance of the gas, diminishes the flow 
and the combustion, and depresses the temperature. 
This depression results in the retraction of the mer- 
cury, a larger inrush of gas, and the temperature 
rises again : thus the regulation takes place con- 
stantly between temperatures so close to each other 
that the oscillations are inappreciable. 

The regulation of the ovens may also be obtained 
by the .expansion of the water between the double 
walls. The water, in heating, expands, rises in a 
small tube, and presses against an elastic membrane 
of caoutchouc or thin metal, which membrane dimin- 
ishes the orifice of entrance of the gas. 

Aerobic cultures. All that is needed in order to ob- 
serve the development of aerobic germs is to place 
the inoculated media in the oven, or, as we may now 
call it, the incubator, at 37; this incubator is so ar- 
ranged that the air in its interior is continually re- 
newed. 

All the media admit of cultures being carried on 
at 37. Nutritive gelatin, however, being fluidified 
above 25, should not be carried above this tempera- 
ture if it is desired to obtain the advantages arising 
from the use of solid media. 



134 Manual of Veterinary Microbiology. 

Anaerobic cultures. Anaerobic germs, in order to 
grow, must be protected from the air ; before attempt- 
ing cultivation, therefore, it is necessary to remove 
the oxygen from the atmosphere and the medium in 
which these are expected to develop. This is accom- 
plished in various ways. The best method consists 
in creating a vacuum in the culture apparatus and 
replacing the air with carbonic acid or hydrogen. 
This operation is repeated several times in order to 
insure the complete removal of the air. Gelatin 
must be fluidified before it is deoxygenated. When 
the operation is completed the culture vessel is sealed 
in the flame. 

The oxygen of the air can be abstracted by sealing 
the culture tube within a second, containing sub- 
stances which rapidly absorb oxygen, such as pyro- 
gallic acid with the addition of a solution of caustic 
potash. Sometimes, also, we add to the culture 
media substances capable of taking up oxygen; for 
example, neutralized sulfate of indigo.* 

* [Anaerobic cultures can also be conveniently carried on in 
small closed tubes completely filled with culture medium. These 
tubes may be filled in various ways; the following method of 
preparation and use is that which I have found most satisfactory: 
A piece of glass tubing about ^ cm. in diameter and 10 to 15 cm. 
in length is sealed at one end in the flame of a blow-pipe, heated 
throughout sufficiently for sterilization, drawn out at the other 
end to a thickness of from 1 to 2 mm., and sealed at a point about 
6 cm. from the thicker part of the tube. The evacuation of the 
air in the tube is obtained by the ebullition of alcohol. A few 
drops are allowed to ascend (by slightly heating the tube, break- 
ing off the point and immersing it in the fluid), shaken to the 
bottom and the tube then held in the flame of a Bunsen burner. 
Long forceps, the points of which are wrapped in asbestos, are 
most suitable for this purpose. The essential points in this process 



Methods of Determination of Pathogenic Microbes. 135 

Physical and chemical characters of cultures. 

Physical characters. When germs proliferate in 
artificial media their cultures assume characters 
which vary with the nature of these germs and with 
the media. 

In cultures on gelatin plates the colonies assume 
different aspects. Sometimes these colonies are rep- 
are that the heating be done carefully in order to avoid breakage 
and that the whole extent of the tube be sufficiently heated to 
prevent condensation of the alcohol vapor. As soon as the alco- 
hol is completely volatilized the open end of the tube is sealed ; an 
abrupt curve near the point insures its breaking at the desired 
place when the tubes are to be filled. A large number of these 
tubes can be made at one time and kept in stock. In making 
bouillon cultures the fluid contained in test-tubes, or (for this 
purpose) better, in small homeopathic vials, is boiled, allowed to 
cool sufficiently, and inoculated with the material from which the 
culture is to be made. The point of the anaerobic tube, after 
quick flaming, is broken off under the fluid by contact with the bot- 
tom of the vial. When the tubes have been properly made, only a 
very small bubble of air should be included with the fluid which 
rushes in. The point is then sealed in the flame, during which 
process a drop or two of the bouillon is necessarily expelled. By 
the use of gelatin or the agar and gelatin mixture, and proper di- 
lution in the usual way, these tubes are quite well adapted for 
isolation of species. The colonies which develop in the substance 
of the solid medium never become large nor show as character- 
istic appearances as by the plate method, but they remain isolated 
and admit of pure cultures being obtained from them. In exam- 
ining with the microscope or making sowings from these colonies, 
I break the tube near the middle, quickly flame one of the seg- 
ments, and carefully heat it at the closed extremity ; the cylinder 
of solid nutrient medium is slowly expelled and received in a 
sterilized Petri dish, from which the " fishing" can be performed 
in the usual way. In tubes prepared in this way strictly anaero- 
bic species can be cultivated, while aerobic species, such as the 
hay bacillus, refuse to grow. D.j 



136 Manual of Veterinary Microbiology. 

resented by small raised droplets of a oily appear- 
ance and of various colors : white, yellow, rose, red, 
purple. In some cases they appear as depressed 
points with regular or sinuous borders; sometimes, 
again, we see a felted mass of filaments radiating 
around a center. Certain germs fluidify the gelatin 
and thus produce a conical depression, full of fluid, 
in the substance of the medium. 

When these colonies are examined under a low 
magnification their surface is seen to be sometimes 
smooth, sometimes granular, their contour regular or 
sinuous, occasionally bristling with filaments. All 
these are peculiarities which serve to distinguish the 
various pathogenic species. 

In gelatin tube-cultures, if -the inoculation has 
been made by puncture through the mass, the inocu- 
lation track may be seen to become turbid and grad- 
ually increase in size; sometimes the germs also 
grow on the surface of the gelatin and spread them- 
selves out in such a way that the whole assumes the 
shape of a nail. "When the growth fluidifies the gel- 
atin around the inoculation track the space which it 
occupies takes the form of a funnel ; but it also hap- 
pens that liquefaction occurs progressively by zones 
from the surface toward the deeper parts. Although 
the inoculation is made in a continuous track the 
growth may only appear in isolated points; this oc- 
curs when only a small number of germs have been 
sown. From the inoculation made into the depth of 
the gelatin a series of tufts may radiate outward, 
thus giving the culture the appearance of a cylindri- 
cal brush or swab. 

Cultures upon the other solid media may present 



Methods of Determination of Pathogenic Microbes. 137 

similar variations. As these media do not become 
fluidified like gelatin, they are better adapted for 
studying the evolution of the germ colonies. 

Some germs give characteristic cultures in certain 
media. Glanders, for instance, upon potato, gives a 
slimy looking culture with chocolate brown margins; 
charbon, sown in gelatin by " stab " inoculation, 
grows in the form of a test tube brush, liquefying 
the nutritive medium. 

Cultures in bouillon are no less variable. Some- 
times we observe a uniform turbidity which is slowly 
deposited, in other cases clots of progressively in- 
creasing dimensions floating in the liquid; sometimes 
a flocculent coherent mass looking like a piece of sat- 
urated wadding suspended in the liquid (charbon) ; 
in still other cases, finally, we see agglomerated colo- 
nies floating on the surface of the bouillon like leaves 
of the water-lily upon water (farcy of cattle). These 
various appearances evidently depend upon the mode 
of association of the germs of which the culture is 
composed. The long filaments take the form of tufts 
of hair; chains of micrococci that of small pellets; 
isolated micrococci produce a uniform turbidity of 
the fluid. 

Germs may change the color of the culture media ; 
such changes are sometimes sufficient to distinguish 
these germs: blue pus. Anaerobic germs often give 
rise to a disengagement of gases more or less offensive, 
recalling those of putrefaction. 

Chemical characters. The chemical reactions to 
which the multiplication of germs gives rise natu- 
rally vary according to the germs concerned; they 
12 



138 Manual of Veterinary Microbiology. 

depend also upon the special conditions in which 
they are placed (aerobic and anaerobic). The medium 
becomes impoverished in alimentary substances, and, 
at the same time, becomes charged with excretory 
products. These we have already described ; we will, 
however, repeat that they may be toxic for the germs 
and arrest their multiplication, and that they often 
communicate to the media in which they are dif- 
fused, certain pathogenic properties which the germs 
themselves possess. 

Preservation of cultures. 

The exhaustion or contamination of the media 
speedily brings the germs to the condition of latent 
life ; from that time they cease to multiply ; the com- 
bined action of the air and light more or less quickly 
destroys them. The virulence of a culture also pro- 
gressively diminishes from the same cause. 

This annihilation of virulence is more or less 
quickly produced according to the germs concerned, 
and also according to the composition of the culture 
medium. Thus, we have seen cultures of fowl chol- 
era deprived of all pathogenic action for adult rab- 
bits after one day, although, usually, this is preserved 
for sixty days, exposed to the air. 

For the longer preservation of cultures of germs 
they may be inclosed in small sterilized tubes which 
are sealed by the flame so as to include as little air as 
possible. These tubes are kept in the dark or in 
wooden cases, as we are in the habit of employing. 
In this way their preservation is lengthened, but it 
is not indefinite; after a variable time, months or 
years, the germs die ; in order to preserve the seed, it 



Methods of Determination of Pathogenic Microbes. 139 

is necessary, from time to time, to invigorate tho 
stock by making inoculations to susceptible animals, 
and from these obtain new cultures. 

III. Experimental infections. 

Pathogenic germs, inoculated to animals, may de- 
termine in the latter troubles of various kinds. These 
artificial microbic diseases are occasionally charac- 
teristic of certain germs. Nevertheless, a given 
germ does not always produce the same morbid con- 
ditions; these are influenced by the animal species 
used for the inoculation and may also vary with the 
individual; the method of inoculation also has its 
influence, and the symptoms observed may be de- 
pendent upon this method ; the condition of the virus 
used as to virulence, and its origin (cultures or 
pathological products), have also an important bear- 
ing on the result of the inoculation ; finally, the 
amount of the virus has an important influence: 
there are diseases which, inoculated in small doses, 
are inoffensive or produce immunity whilst large 
doses more or less rapidly result in death. 

All these reasons indicate that we can draw no ab- 
solute conclusions as to the results of the inoculation, 
and that the latter is of no value except when com- 
bined with other means of diagnosis. 

Experimental inoculations are occasionally made 
with the object of purifying an impure virus ; in this 
case advantage is taken of the property possessed by 
certain germs of developing in a given animal 
species, whilst the other microbes with which they 
may be commingled can not live in this species. 

Subjects of inoculation. Many species of animals 



140 Manual of Veterinary Microbiology. 

may, in an emergency, be brought into use for experi- 
mental inoculation, but there are some which are pre- 
ferred on account of their great susceptibility for most 
of the bacterial diseases and of the ease with which 
they can be obtained. 

The animals which are the most used are the 
guinea pig, rabbit, rat, mouse, chicken, pigeon, and 
small birds. In exceptional cases the large domes- 
ticated animals are employed. 

Inoculation substances. Virulent substances used 
for inoculation should, as far as possible, be free from 
all microbic mixture, and the Pravaz syringe and 
other inoculation instruments should previously be 
rendered aseptic. 

If the product to be inoculated is a bouillon culture, 
this fluid is turned, after shaking, into a previously 
flamed watch glass, and quickly aspirated into the 
syringe. 

If the culture has been made upon a solid medium 
a quantity of -the material is taken upon a sterilized 
platinum wire and diluted in a little sterilized water 
or bouillon. 

When we have to do with fluid pathological pro- 
ducts (blood, milk, pus, etc.) these products, col- 
lected in a pure state from the living being or from 
the cadaver, are employed in the way just described. 
In some cases they are previously diluted. 

Sometimes the virulent substance is a solid patho- 
logical product. This product, free from all con- 
tamination, is crushed in a special mortar, diluted in 
bouillon, which is then strained through fine linen. 
To more completely avoid the external germs the 
virulent particle may be introduced into a small 



Methods of Determination of Pathogenic Microbes. 141 

previously sterilized tube, and crushed by means of 
a flamed glass rod, the diameter of which is a little 
smaller than that of the interior of the tube ; steril- 
ized bouillon is then added, and, after the particles in 
suspension have been deposited, it is aspirated into 
the syringe. 

Methods of inoculation. The virus, thus prepared, 
may be used for inoculations performed in various 
ways, which we will now review. 

1. Endermic inoculations. This is the simplest 
method of inoculation ; in its performance all that is 
necessary is to lay bare the deeper layer of the epi- 
dermis without exciting much hemorrhage, and there 
apply the active substance. 

The hair is clipped from the region to be inocu- 
lated, a series of closely approximated superficial 
scarifications made with a bistoury, and the virulent 
substance spread over it. For this inoculation regions 
of the body should be selected which are not easily 
reached by licking, rubbing, etc. 

2. Subcutaneous inoculation. The object of this in- 
oculation is to introduce the active product into the 
subcutaneous cellular tissue. A fold of the skin hav- 
ing been pinched up, the canula of the syringe is in- 
troduced at its base and the liquid to be inoculated 
slowly expelled. It is sometimes desirable to free 
the region from hair or feathers and render the point 
of inoculation aseptic, by the application of a strongly 
heated glass rod. The inoculation having been made, 
the canula is withdrawn, and, in order to render the 
result more certain and more rapid, the inoculated 
point is manipulated so as to lacerate the cellular tis- 
sue and accelerate absorption. 



142 Manual of Veterinary Microbiology. 

Inoculations may be made in any region, but, 
preferably, in places where the skin is thin and pliant 
(internal face of the thighs, abdominal wall, pectoral 
region in birds), and the cellular tissue abundant. 

3. Intra-peritoneal inoculations. In order to intro- 
duqe the virulent matter into the abdomen, the ani- 
mal being held well on the back, the abdominal walls 
are pinched up between the thumb and index, and 
the operation continued as above, after being well as- 
sured that the point of the syringe is indeed free 
within the abdomen. It is necessary here to operate 
with a certain amount of care in order to avoid 
wounding the viscera an accident especially to be 
feared in birds. 

4. Intra-venous inoculation. This method necessi- 
tates the employment of a liquid free from solid par- 
ticles which, by their arrest in the smaller vessels, 
might occasion fatal embolisms. In performing this 
inoculation the vein is distended by pressure exerted 
on its course and the canula introduced into it, the 
point directed toward the heart ; the operation is suc- 
cessful when a drop of blood issues at the shoulder 
of the canula; a graduated pressure is then brought 
to bear on the piston. 

For this inoculation the most salient superficial 
veins are selected, in the rabbit the veins of the ear, 
in birds the vein of the arm. In some animals the in- 
oculation can be made into the jugular, saphena, etc. 

5. Inoculations in the anterior chamber of the eye. 
The eye is first ansesthized by means of a few drops 
of a solution of cocaine, at 1 to 20; then, the globe 
being immobilized, the canula is insinuated horizon- 
tally through the cornea in its eccentric part. One 



Methods of Determination of Pathogenic Microbes. 143 

or two drops of the liquid are then expelled and 
may be seen diffusing themselves in the transparent 
media. 

. 6. Iktra-cranial inoculations. It is necessary here, 
in the first place, to trephine the cranium, an opera- 
tion which necessitates a variety of instruments. In 
small animals, however, the operation can be very 
simply performed. The animal being made fast, an 
incision is made in the skin over the frontal bone out- 
side of the median line, the periosteum is crucially 
divided and, with the point of a strong bistoury held 
vertical to the surface of the bone, a small opening is 
made by rotating the instrument upon itself. 

When the opening is considered to be of sufficient 
size the canula, thoroughly sterilized, is introduced 
under the cerebral envelopes and a few drops of the 
liquid injected. 

Inoculations may also be made into the various 
serous cavities, the trachea, muscles, etc., but the 
technique of these operations needs no special de- 
scription. 

Besides these contagions by inoculation, an organ- 
ism may also be artificially infected by the methods 
of ingestion or inhalation of virulent products. These 
methods give results relatively less certain. 

Collection of virulent >rodi/ds. Pathological prod- 
ucts can be collected either from diseased animals or 
from their cadavers. 

On diseased animals the process differs according as 
it is desired to collect a liquid (blood, pus, etc.) or a 
solid particle. Great precautions should always be 
taken in order to avoid the common germs which 
surround us. After the region has been shaved it 



144 Manual of Veterinary Microbiology. 

must be sterilized by means of a strongly heated 
glass rod. The solid part which it is desired to study 
is then extracted with flamed instruments; liquids 
are aspirated into the Pasteur pipette. 

On cadavers the same precautions are necessary. It 
must be remembered that, in contagious diseases, de- 
composition usually proceeds very rapidly and, in 
consequence, the organism soon becomes invaded by 
the germs of putrefaction. The cadaver having been 
fixed in the proper position, the autopsy must be con- 
ducted in a methodical manner and the products, 
solid or liquid, which it is desired to study must be 
collected aseptically. 

It should not be forgotten that the disease in ques- 
tion is a contagious one, and that no part of the ani- 
mal should escape the destruction which ought to 
follow all autopsies of this kind; it is well, also, to 
remark that several microbic diseases are transmissi- 
ble to man ; the operator, therefore, should take the 
precaution to -protect himself against infection. 

Solid products, when collected, should immediately 
be used for inoculations and for cultures, or should be 
inclosed in sterilized tubes. Liquids should be sealed 
in the Pasteur pipettes in which they have been col- 
lected. 



Microbic Diseases Individually Considered. 145 



PART THIRD. 

MICROBIC DISEASES INDIVIDUALLY CONSID- 
ERED. 



Microbic diseases consecutive to wounds. 

Surgical wounds, treated strictly according to anti- 
septic rules, are protected from pathogenic germs ; 
they heal without excessive swelling and without 
suppuration or fever. The healing process is limited 
to the extent of cell proliferation necessary for repa- 
ration only. It is the same with accidental wounds 
when they are rendered aseptic ; but if, by the con- 
tact or the subsequent action of the object causing 
the injury, or by contact with clothing of any kind, 
harness, litter, water, or the atmosphere, pathogenic 
germs are introduced into the wound, diverse patho- 
logical changes may be observed. 

We have seen, in the general part of this work, 
that a number of pathogenic germs, for example, 
those of suppuration and septicaemias are encountered 
almost everywhere; we meet with them also in most 
of the morbid conditions which complicate wounds ; 
these diseases we shall first of all consider. 

The lesions of a microbic nature which develop 
consecutive to wounds are local, remote, or general. 
13 



146 Manual of Veterinary Microbiology. 

Local lesions consist of inflammatory processes the" 
predominant character of which depends principally 
on the special pathogenic property of the microbes 
contained in the wound; these processes may be es- 
sentially exudative ; inflammatory cedemas, erysipelas; or 
suppurative : superficial suppuration, abscess, phlegmon; 
or hypertrophic : actino'mycomata, botryomycomata, an- 
atomical tubercles: or, finally, gangrenous: traumatic 
gangrene, diphtheria, hospital gangrene. The dominant 
character of these inflammations depends principally, 
but not exclusively, on the special nature of the germs 
distributed upon the wounds; in reality, the reaction- 
ary powers of the tissues have also their influence 
here, and a given germ, the streptococcus of erysipe- 
las for example, will occasion sometimes a simple 
dermatitis with interstitial and superficial exudation 
under the form of vesicles, phlyctenae (erysipelas), 
sometimes it will determine, in addition, a deep seated 
suppuration (phlegmonous erysipelas), and even ne- 
crosis of the- inflamed tissues (gangrenous erysipelas). 

Remote lesions manifest themselves in the organs 
in direct or vascular continuity with the tissues orig- 
inally attacked. Hence, we see peritonitis occur 
consecutive to changes of a microbic nature located 
in the abdominal viscera : wounds of the intestine, 
of the uterus, metritis, etc. In respect to vascular 
continuity, this involves alterations of the correspond- 
ing lymphatics and blood vessels (lymphangitis, adenitis, 
phlebitis, endocarditis, thrombic and embolic lesions). 

Finally, as general troubles, we have pycemia and 
surgical septicaemias. 



Microbic Diseases Individually Considered. 147 

Suppuration. 

We find in pus, besides the cells which constitute 
its essential part, various species of microbes. Their 
constant presence in suppurative inflammations led to 
the supposition that suppuration was only produced 
through the agency of microbes. This theory was 
further supported by the results of experimentation ; 
thus, subcutaneous injection of irritating substances 
previously rendered aseptic caused an inflammation 
corresponding to the irritating power of these sub- 
stances, but a non-purulent inflammation. Diapede- 
sis of the white corpuscles of the blood seemed there- 
fore to be dependent upon the presence of microbes. 

Similar experiments, however, have led to contrary 
results 'in the hands of other investigators, and sup- 
puration has been produced by means of chemical 
substances (croton oil, oil of turpentine, silver nitrate, 
mercury, cadaverin, etc.) without the intervention of 
bacteria. Moreover, it has been found that the ster- 
ilized cultures of the staphylococcus pyogenes as 
surely determine an abscess as the staphylococcus 
itself; the pus of this abscess, however, is not itself 
pyogenic. Substances with special pyogenic proper- 
ties have, in addition, been extracted from cultures 
of the bacilli of glanders, tuberculosis, charbon and 
of Friedlander's pneumococcus. Thus is explained 
the possibility of spontaneous abscesses the pus of 
which is free from taiicrobes ; the latter, in such case, 
are present in another part of the economy and the 
pyogenic substances which they secrete being ab- 
sorbed, we can understand that they may determine 
purulent inflammation of a predisposed organ or 
tissue. 



148 Manual of Veterinary Microbiology. 

These considerations, important as they are from a 
theoretical point of view, have, in practice, only a 
secondary interest. Under natural conditions suppur- 
ation is really always the result of microbes, acting 
not of themselves, as was at first supposed, but, ac- 
cording to recent researches, by means of the toxines 
to which their nutritive exchanges give rise.* 

The rational application of antiseptics in the dress- 
ing of wounds has, further, to a large extent, demon- 
strated this truth, in making union by first intention 
the necessary termination of operative wounds. 

The most common bacteria of suppuration are : 
the staphylococcus pyogenes aureus (yellow pus) ; 
the staphylococcus pyogenes albus (white pus) ; 
the staphylococcus pyogenes citreus; 
the streptococcus pyogenes, 

and several other species, including a bacillus the 
bacillus pyogenes septicus. 

These are met with in the various suppurative 
processes: phlegmons, abscesses and the effusions of 
pyaemia, purulent inflammations of the external and 
internal surfaces, etc. 

The yellow staphylococcus has also been found in 
furuncles and malignant pustule; the particular char- 
acters of these diseases seem to depend on the mode 
of penetration and the localization of the pyogenic 
germs rather than on the special nature of the latter. 
We have ourselves encountered the staphylococcus 
albus in furuncles which had developed in large num- 

* [According to Buchner's investigations, the pyogenic property 
of sterilized cultures of many bacterial species resides in the bac- 
teria themselves, and not in the chemical products which they 
secrete. D.] 



Microbic Diseases Individually Considered. 149 



Fig. 2. 



bers on the back of a colt a few weeks old, and 
which was successfully treated in the college hos- 
pital. 

The streptococcus pyogenes seems to be also the 
causative agent in erysipelas and puerperal fever under 
its various forms ; at least, the streptococcus of these 
two diseases can not be satisfactorily differentiated 
from that which we are now considering. 

The streptocooccus pyo- 
genes is widely distributed 
and its virulence is subject 
to great variations; when 
injected to animals, and es- 
pecially to the rabbit, it 
most frequently gives rise 
to a local abscess, but it can 
also, like the staphylococcus 
pyogenes, occasion multiple 
abscesses in the muscles, 
kidney, lung, etc., and more 
or less quickly bring about abscess of horee ' X i.OOO.-D. 
a fatal termination. The staphylococcus aureus is 
encountered still more frequently than the strepto- 
coccus. 

Cultures. The pyogenic germs are easily culti- 
vated in the different media ; the three staphylococci 
are distinguished from each other by the color of 
their cultures, the aureus giving a golden yellow 
color, the albus a white color, and the citreus a citron 
yellow color. 

The staphylococci pyogenes are aerobic ; the strep- 
tococcus is rather anaerobic. 

The chemical study of cultures of the staphylococ- 




Streptococci in pus of closed 



150 Manual of Veterinary Microbiology. 

cus aureus has led to the discovery of a non-toxic pto- 
maine, a non-nitrogenous phlogogenic and pyogenic 
substance called phioyosine, and a diastase also pyoge- 
nic. The yellow coloring matter is only developed in 
contact with the air. 

Research and coloration. In order to bring into 
view the microbes of pus it suffices to stain the latter 
on cover glasses for a few seconds with a hydro-alco- 
holic solution of gentian violet. The Gram reaction, 
which they all sustain, admits of obtaining double 
coloration. "We, by this means, see the microbes be- 
tween, and often even inside, the pus corpusles. 

Etiology. The germs of pus most frequently pene- 
trate into the tissues through a solution of continuity, 
but they may also invade the organism when the 
skin is intact; this has been demonstrated by Garre; 
frictioning of the arm with a culture of staphylococcus 
aureus caused, on the same day, the eruption of 
furuncular pustules. Numerous examples of this 
truth are furnished by veterinary accoucheurs, in 
whom the arm becomes covered with pustules as a 
result of their intervention in difficult parturition 
or in removal of the placenta. In this case the le- 
sions always develop at a pilo-sebaceous follicle, 
through the orifice of which the purulent germs have 
entered. Finally, the germs of pus may gain the 
tissues by traversing the normal mucosa. 

The pyogenic germs which we have just referred 
to are not the only ones which can give rise to the 
formation of pus. A number of others possessing 
specific virulent properties have, in addition, the 
power of exciting suppuration ; among these are the 
Koch bacillus, the bacillus of glanders, the actino- 



Microbic Diseases Individually Considered. 151 

myces, the bacillus of cattle farcy, the bacillus of 
chicken cholera in the guinea pig, the bacillus of 
typhoid fever, etc. These bacteria, incidentally pyo- 
genic, have been qualified as pyocolic ; they act by 
themselves or by preparing the field for the pyogenic 
germs properly so called. It may be further stated, 
in this connection, that the mere contact of pyogenic 
microbes with a wound or living tissue is not invari- 
ably followed by suppuration. Here, the number of 
microbes as well as the resistance of the tissue or of 
the organism to which the latter belongs, all enter 
into the question. It is a matter of common obser- 
vation that even a simple wound in some individuals, 
if left to itself, will always come to suppuration. The 
adjuvant action of traumatisms, irritant liquids, and 
individual predisposition has, here, an important 
bearing, modifying the resistance of the tissues toward 
pyogenic germs. 

The action of pyogenic germs can be experiment- 
ally aided by the injection of their culture products. 
Thus, the filtered cultures of the staphylococcus and 
streptococcus pyogenes contain adjuvant substances 
and vaccinating substances. The first are the most 
active and alone show their effects when these filtered 
cultures are injected; but they are destroyed by 
twenty-four hours exposure to 55 in the case of the 
staphylococcus, and to 110-for the streptococcus, and 
the injection of cultures thus heated confers immunity. 
According to Courrnont and Dor, the adjuvant action 
of filtered cultures of the streptococcus endures for 
at least three months. 

Blue pus Pus, in the human being, occasionally 
exhibits a peculiar blue color ; this is the result of a 



152 Manual of Veterinary Microbiology. 

special polymorphic microbe, generally assuming the 
form of a short curved bacillus, occasionally that of 
a micrococcus or even of a spirillum. 

The germ of blue pus is very easily cultivated ; it 
communicates to the nutritive media a green color ; 
by the action of chloroform it is possible to isolate 
form its cultures pyocyanine which is the blue color- 
ing matter characteristic of the germ. The green 
color of the cultures results from the fact that the 
nutritive media have originally a yellowish color. 

When injected to animals it does not cause suppu- 
ration ; but its cultures are, nevertheless, pathogenic 
for the rabbit to which it gives a special disease, the 
pyocyanic disease, acute or chronic according to the 
dose employed, and characterized by paralysis, fever, 
albuminuria, and diarrhoea. 

The microbe of blue pus has been encountered in 
our animals only by M. Cadeac, in the spleen and 
lymph glands of a dog killed in the last stage of 
lymphadenoma. The author tested its identity by 
culture methods and inoculations but did not succeed 
in transmitting the disease to the dog ; he thinks that 
the abnormal debility of the lymphadenomic subject 
facilitated the installation of the germ of the pyocy- 
anic disease. 

Pycemia. 

The knowledge which we now possess of recep- 
tivity and the conditions which determine it, as well 
as of the situation of the pyogenic germs in the 
meshes of the connective tissue in cases of phlegmon, 
justifies us in asserting that the passage of these 
germs into the circulatory fluids is probably a com- 
mon but generally inoffensive occurrence. If the 



Microbic Diseases Individually Considered. 153 

organism should suffer from any serious disturbance, 
or if, in consequence of a special local alteration, these 
germs should penetrate in large numbers into the 
blood, their influence will cease to remain limited to 
their original focus and we will see the irruption of 
that formidable disease, pyaemia. This disease is not 
absolutely dependent upon the existence of a wound; 
it can originate in the course of a purulent visceral 
inflammation or even appear spontaneously. In this 
last case the microbes come either from the mucous 
membrane or from an old latent focus. 

Besides the immediate general phenomena, such as 
those of a peculiarly intense fever, the arrest and 
multiplication of the pyogenic germs in different 
parts of the circulatory apparatus lead to the produc- 
tion of metastatic abscesses and purulent collections 
in the natural cavities. The pyogenic germs com- 
municate to the blood corpuscles a certain degree of 
viscidity; the latter become agglutinated and, as a 
result, embolisms occur in the small arterioles of one 
or more organs : kidney, liver, lungs, muscles, etc. 
The microbes arrested at these embolisms form the 
starting point of so many foci of suppuration. 

In reality pyaemia is rarely simple; along with 
the bacteria of pus the blood generally receives other 
microbes from wounds exposed to the air, and there 
results a concomitant disease of a septicsemic order. 
We shall see, further, that the organisms of pus may, 
under certain circumstances, give, of themselves, a 
pure septicaemia. 

Septiccemia. 

In a general way this name is given to the patho- 
logical condition which follows penetration of putrid 



154 Manual of Veterinary Microbiology. 

matters into the system. We now know, however, 
that putrefaction is only a complex fermentation 
tending to the degradation of organic matter, with 
or without the production of fetid odor, a process 
which necessitates the intervention of germs or mi- 
crobes of various species. Hence we can not con- 
ceive of septicaemia without microbes. 

But the r6le of the latter has not always the same 
importance ; the germs, remaining intrenched outside 
the tissues upon a wound, for example, act solely 
through the soluble products (ptomaines and dias- 
tases) to which their nutritive exchanges give origin, 
and we then have to do with a kind of poisoning ; or, 
the same germs, breaking the barrier which the liv- 
ing tissues oppose to them, make their way into the 
latter and multiply there; intoxication by toxines 
then becomes complicated with troubles caused by 
this proliferation. 

The first of , the two contingencies which we have 
just mentioned occurs under a multitude of condi- 
tions. And, first of all, it is unquestionable that sep- 
ticaemia sometimes can be produced without the pres- 
ence of microbes. Koch has shown that five drops 
of a putrefying fluid kills a mouse in a few hours, no 
microbes then being found either in the blood or vis- 
cera. The disease thus developed is a true intoxica- 
tion by soluble microbic substances or ptomaines en- 
gendered outside of the organism in the putrefying 
fluid. It is to this species of septicaemia that we 
must refer the intoxications which result in animals 
and man from the consumption of imperfectly pre- 
served foods in way of decomposition or already 
decomposed; such, for example, as the poisoning 



Microbic Diseases Individually Considered. 155 

(botulisme) which occurs in the human species from 
the use of so-called Boulogne sausage prepared by 
unscrupulous dealers from the flesh of animals which 
have died or been slaughtered, diseased or sound, but 
nearly always, under the mask of seasoning, already 
abandoned to the sway of microscopic life which not 
only uses up its substantial parts, but elaborates di- 
verse ptomaines of formidable toxic power. 

At other times septicaemia is a sort of auto-intoxi- 
cation, the septic poisons being elaborated within the 
economy itself but upon a limited surface which the 
microbes do not break through, either by reason of 
their special properties (the anaerobes are unable to 
live in the blood during life), or because the toxicity 
of the products secreted and absorbed is such that 
death supervenes in too short a time to allow of 
the invasion of the circulatory fluids ; this surface is 
most frequently the seat of a morbid microbic pro- 
cess : for example, a septic wound, puerperal metritis, 
gangrenous pneumonia, etc. 

But the germs of the intestine may give rise to 
troubles even in the absence of special alterations of 
the mucosa. This takes place when their products of 
denutrition ptomaines, indol, skatol, gaseous prod- 
ucts, etc. instead of being eliminated by way of the 
rectum, are absorbed by the blood. The chemical 
poisoning which then results has received the names 
stercorcemia and intestinal septicaemia. This poison- 
ing supervenes under a number of pathological con- 
ditions, especially when the gastric juice is insuffi- 
cient in quantity or in acidity to neutralize the ma- 
jority of the germs which pass through the stomach, 



156 Manual of Veterinary Microbiology. 

and, in a general manner, in all somewhat lengthened 
diseases which lead to a prolonged retention of fgecal 
matters in the intestine (fever, inappetence, retention 
of bile, etc.). The indication, in these conditions, 
is to retard the intestinal fermentations by the ad- 
ministration of special antiseptics, and to assist in 
the evacuation of the toxic substances and the germs 
which produce them, by means of purgatives. This 
evacuant action of laxatives explains their utility in 
all febrile affections in which, for special reasons, 
these remedies are not contra-indicated. 

,The pathogeny of septicaemia, therefore, is not of 
one kind only. The nosological extent of the term 
septicaemia is, moreover, very imperfectly defined. 
When we take into account the fact that bacteria 
contribute in a general way to the reduction of com- 
plex organic molecules to formulae more and more 
simple, and, on the other hand, consider the difficulty 
of determining, in this immense work of microbes, 
what should be regarded as putrefaction, and what 
definition should be given of putrid matters, we must 
recognize that, from the mode of action of the germs 
which occasion them, as well as by their evolution, 
all general diseases of a microbic order ought to be 
included in the group of septicaemic affections. If a 
certain number of these have been placed apart, this 
is on account of the specific characters of their germ 
which, giving to the disease a special expression, have 
established for it a well defined morbid entity : char- 
bon, typhoid fever, tetanus, etc. 

Prof. Degive had already twenty years ago con- 



Microbic Diseases Individually Considered. 157 

ceived and clearly expressed this manner of view in 
several reports and discussions. (1) 

He recognized in these diseases actual points of 
consanguinity which led him to refer them all to -the 
same stock and to regard them as children of the 
same family, " The septoid stock or family, also called 
septicaemia, but better designated septose, the septic or 
putrid diathesis." If the discovery of the special 
germs of most of these diseases no more allows of 
belief in their identity, it removes all doubts as to 
their analogy. 

The name saproemia has been reserved for septicae- 
mias engendered by microbes which develop a strong 
odor of putrefaction. 

Septicaemia was formerly considered only as a com- 
plication of wounds, but it can occur independent of 
all traumatism, the agents which occasion it pene- 
trating by one of the natural surfaces without the aid 
of any solution of continuity, as, for instance, by the 
digestive, respiratory, genital, urinary passages, etc. 
There are, therefore, surgical septicaemias and medical 
septicaemias. 

When septicaemia develops in consequence of a 
traumatism we see in the latter important changes 
supervene which indicate the infection of the wound 
by germs : the secretion becomes sero-sanguinolent, 
often fetid, the granulations become less firm, pulta- 
ceous, often purple; the neighboring tissues swell 
and become the seat of an inflammatory oadema, fre- 
quently progressive ; these local lesions may entail 

(1) See Annales de medicine Veterinaire, 1874, p. 502; 1875, p, 
94, and 1876, p. 115. 



158 Manual of Veterinary Microbiology. 

very serious disorders, such as gangrene and detach- 
ment of the adjoining tissues; sometimes an actual 
putrefaction establishes itself in the living animal on 
the part invaded by the germs. We can not, how- 
ever, assign any fixed rule as to the importance of the 
local troubles ; they manifestly depend upon the na- 
ture of the germs which occasion them. 

As to the general symptoms, they are the conse- 
quence of the absorption of the toxic substances 
elaborated by the micro-organisms at the wound it- 
self. The appearance of febrile symptoms, and more 
especially abnormal elevation of temperature, are the 
first indications of the intoxication ; they enable the 
practitioner, in the absence of any other evident 
cause, to assure himself of the infection of the wound, 
and they constitute an important indication for its 
subsequent treatment. Along with this, poisoning 
of the nerve centers by the bacterial ptomaines 
shows itself by the phenomena of coma, stupor, or 
even delirium. 

The blood -corpuscles, in contact with these sub- 
stances, undergo a more rapid destruction, and their 
coloring matter, in excess in the plasma, communi- 
cates to the interstitial fluids, and notably to the 
visible mucous membranes, a dull yellow tint more or 
less intense, which, in severe cases indicates the ex- 
istence of a kind of hsemaphic icterus ; the produc- 
tion of this last condition is facilitated by the circum- 
stance that the liver, itself altered, is unable to 
eliminate the coloring matter which the blood plasma 
carries to it in excessive amount. 

The germs of septicaemias are not always found in 
the blood; some of them are anaerobic and unable to 



Microbic Diseases Individually Considered. 159 

multiply there ; in the case of others they may be 
seen to penetrate into the circulatory fluids when the 
disease is of rather long duration. It is evident that 
this penetration can be followed by new troubles 
localized in the different parts of the economy in 
which these germs become arrested. But even in the 
absence of microbes in the circulation, general symp- 
toms of septicaemias are nearly always complicated 
with local symptoms due to the irritant action of the 
absorbed ptomaines. Such is the pathogeny of 
parenchymatous nephritis with albuminuria, hepatitis, 
enteritis with diarrhoea, pneumonia, pleurisy, peri- 
carditis, myocarditis, endocarditis, meningo-enceph- 
alitis, etc., which are seen to develop in the course 
of septicaemias. 

Septicsemic fever presents several varieties, the 
clinical importance of which should not be disre- 
garded. Traumatic fever is the mildest form, the first 
degree of acute septicaemia ; the latter develops in sev- 
eral days and determines a greater and more lasting 
elevation of temperature; super-acute, fulminating, or 
gangrenous septicaemia kills the subject in a very short 
time ; finally, chronic septiccemia, or hectic fever occa- 
sions a slow pining away of the patient, and is of 
much longer duration. 

Septicaemia may succeed to a large number of local 
lesions : sanious wounds, abscesses, erysipelas, fu- 
runcle, gaseous gangrene, etc. The microbes which 
occasion it are very variable and the symptoms are 
therefore not always alike. 

The pyogenic germs can also give rise to a pure 
septicaemia; the streptococcus pyogenes is the cause 
of puerperal fever; the staphylococcus pyogenes 



160 Manual of Veterinary Microbiology. 

aureus has been met with in the blood in several 
cases of septicaemia. In such cases the virulence of 
these germs is generally very great and death super- 
venes too quickly to allow of the formation of pus. 
We thus see puerperal fever under three different 
types in which the streptococcus pyogenes is always 
found; it sometimes assumes the form of a true sep- 
ticaemia quickly leading to death ; at other times the 
patient succumbs with an abscess of the large liga- 
ments and generalization of the streptococcus, with- 
out occasioning new abscesses; finally, the disease 
sometimes evolves comparatively slowly, assuming 
the characters of a pyaemia with multiple strepto- 
coccus abscesses. 

These observations show that in septicaemia and 
pyaemia the process is essentially the same ; the re- 
sult, in case of pyaemia, proceeds from the special 
pyogenic property of the germ and from its par- 
ticular degree of virulence. 

The lesions found at autopsies of septicaemic sub- 
jects are far from being constant. However, it is ob- 
served that the bodies rapidly putrefy. The paren- 
chyma of the liver, kidney, and spleen are often in- 
flamed and softened. We may also find inflammation 
of the various serous membranes: pleura, peri- 
toneum, pericardium, endocardium, etc.; multiple 
hemorrhages and the more or less icteric color of 
all the tissues indicate, in certain cases, profound 
alteration of the blood. 

Pasteur's septicaemia. 

We will not delay to describe the history of the 
septicaemias experimentally obtained; they present, 



Microbic Diseases Individually Considered. 161 

indeed, great interest from a general bacteriological 
point of view, but, here, we will only study those 
germs which the veterinarian is liable to meet with 
in practice and which it is absolutely necessary that 
he should be acquainted with. 

The bacillus septicus or septic vibrio takes first rank 
among those which give rise to the most character- 
istic phenomena of septicaemia. It is the cause of the 
complications of wounds described under the names 
of gaseous gangrene, fulminating gangrene, traumatic 
gangrene, and malignant oedema. The disease which 
it occasions in man and animals has been desig- 
nated gangrenous septicaemia by MM. Chauveau and 
Arloing. 

Characters of the septic vibrio. It is a rod measuring 
ty in length by I// in breadth, hence shorter than 
that of bacteridian charbon; it is often jointed like 
the latter but its different segments 
have not all the same length, whilst 
the articulated segments of char- 
bon are of uniform dimensions. 
Further, whilst the segmented bac- 
teridia are cut at right angles and 
slightly swollen at their ends, these 
characters are not observed in the 

septic bacillus. According to Chau- ^ *> 2 > , 3 ' Bacilli of 

, . , . , . , Pasteur s septicaemia 

veau and Arloing, when examined in the peritoneum 

in the oaderna of a septicsemic focus, and in the blood, 
it shows itself: 1st. with the char- 4, 5. Segmented 

acters of a bacillis (6/z to 50/* by I// to bacilli - 

_ N . i i j. 6. Bacillus with 

1.5/) provided with a spore at one terminal Bpore ._ M . 

of its extremities, which is occa- and L. 
24 




162 Manual of Veterinary Microbiology. 

sionally swollen ; 2d. or those of a bacillus with 
homogeneous protoplasm, a little longer than the 
preceding (12/z to 30//). In the serous membranes, 
and in the blood after death, it grows to a consider- 
able length and more or less rapidly becomes seg- 
mented into articles of varying lengths, never sporu- 
lated after the manner of the bacillus of charbon. 

The septic vibrio is absolutely anaerobic ; it exhibits 
very active flexuous movements which are quickly 
arrested by contact with oxygen. 

Action of physical and chemical agents. Heat is the 
surest and most active agent of destruction of the 
septic vibrio and the only one to be recommended in 
practice; virulent serosity is rendered inoffensive by 
heating for fifteen minutes at 100; dried serosity re- 
quires a little less than ten minutes at 120. It slowly 
loses its virulence through the influence of putrefac- 
tion (in two months) ; the virus, dried at temperatures 
varying from 15 to 38, is indefinitely preserved. 
Antiseptics have a feeble toxic power for the septic 
vibrio; according to MM. Chauveau and Arloing sul- 
phurous acid has shown itself the most powerful; 
sublimate, at 1 to 500, does not kill the bacillus of 
gangrenous septicaemia. Carbolic acid, at three 
per cent, is only efficacious when supplemented by 
heat. 

Cultures. The bacillus septicus multiplies in all 
the artificial media under the express condition that 
these media and the atmosphere in which they are 
inclosed are deprived of oxygen. The development 
is accompanied with the disengagement of carbonic 
acid and hydrogen ; bouillons become turbid and then 
clear by the deposition of the bacilli ; gelatin is fluid- 



Microbic Diseases Individually Considered. 163 

ified. Agar is well adapted to its culture ; pn this me- 
dium it forms a whitish track with festooned borders 
and extends through all the nutritive mass through 
the breaking up of the latter by the liberated gases. 
In bouillon the bacilli are undulated or straight; at 
first homogeneous, they later become granular and 
break up. Some of the elements become sporulated; 
the spore most frequently appears at a swollen ex- 
tremity of the bacillus and gives to the latter the ap- 
pearance of a bell clapper. When cultivated upon 
solid media the bacilli are shorter, and fructification 
is more delayed. 

Inoculation of culture media should be made either 
from the peritoneal serosity or from the blood of 
septicsemic subjects. But as the blood contains very 
few germs immediately after death, it is necessary, 
for the inoculation to be successful, to allow them to 
multiply there. For this end a little blood is inclosed 
in a pipette and kept in the incubator for twenty-four 
hours ; multiplication of the bacilli takes place and 
soon shows itself by the appearance of bubbles of 
gas. The blood may then be employed for the in- 
oculation of artificial media. Muscle juice can be 
used for the same purpose. 

Research and coloration. Examination should be 
made of the serosity of the oedema or of the septi- 
csemic focus, of the blood, peritoneal serosity, and 
muscle juice. By studying fresh unstained prepara- 
tions we can appreciate the movements of the bacilli 
which wind among the elements of the blood or se- 
rosity ; it will be noticed that the rods in the center of 
the preparation preserve their motility much longer 



164 Manual of Veterinary Microbiology. 

than those in the vicinity of the margin of the cover 
glass where they are killed by contact with the air. 

Experimental inoculations. The species endowed 
with receptivity for the bacillus of gangrenous sep- 
ticsemia are : guinea pig, rabbit, sheep, goat, horse ; then 
in the order of decreasing affinity, the ass, chicken, 
pigeon, and, finally, the dog and cat. Cattle are abso- 
lutely refractory ; this is a remarkable exception which, 
itself, is sufficient to differentiate this pathogenic 
agent from that of symptomatic charbon, with which 
is has several points of resemblance. 

Inoculation is most successful when made in the 
subcutaneous cellular tissue (one-fifth of a drop to 
five drops of the virulent serosity are sufficient) ; it 
invariably fails when made with the lancet or by su- 
perficial scarifications. When injected into the cir- 
culation the animal can tolerate doses much larger 
than those which prove fatal when injected into the 
subcutaneous cellular tissue. Subjects inoculated in 
this way suffer from a fever of more or less intensity 
and obtain immunity; but, when the quantity intro- 
duced into the blood has surpassed one to three drops 
of the virulent serosity in the rabbit, one to five cubic 
centimeters in the sheep, ten to thirty-five cubic centi- 
meters in the ass, it leads to the death of the inocu- 
lated subject. In animals which have been rendered 
refractory by intra- vascular injection later inocula- 
tion in the connective tissue produces a slight swell- 
ing or, at most, the formation of a curable abscess. 

The serosity taken from the muscular tissue, the 
connective tissue, and the parenchymatous organs, is 
more virulent than that from the serous cavities. 

Inoculation in the connective tissue is followed by 



Microbic Diseases Individually Considered. 165 

a violent inflammatory reaction ; the region becomes 
hot, painful and tumefied ; the engorgement rapidly 
extends to neighboring regions and becomes crepitant, 
emphysematous, in consequence of the internal forma- 
tion of gas (carbonic acid, hydrogen, carburetted and 
sulfuretted hydrogen, etc.). Soon the central part 
becomes insensible and presents all the symptoms of 
mortification. Gaseous infiltration may be wanting 
when death supervenes too quickly. The latter ar- 
rives at the end of twelve to fifteen hours in the 
guinea pig. The autopsy shows great detachment of 
the tissues, their infiltration with a sanious fluid and 
fetid gases, gangrenous patches of greater or less ex- 
tent, and often great extension in all directions of the 
original inflammation. 

Intra-vascular inoculation may be followed by the 
same changes when a solution of continuity of the 
circulatory apparatus allows the germs to penetrate 
into the connective tissue and multiply there, shel- 
tered from the oxygen of the blood. 

The disease is transmissible from the mother to the 
foatus. 

Etiology and pathogeny. The septic vibrio is almost 
every- where present in the soil, the dust of hay, in 
most putrid substances and also in the digestive canal 
of healthy animals. But in this last situation it is 
inoffensive; after death, however, the oxygen be- 
coming deficient in the tissues which during life op- 
pose themselves to its penetration, it invades these 
tissues, multiplies there, and in this way we . are able 
to establish its presence in the blood (first of the 
portal vein, then throughout the economy) and on 
the surface of the abdominal viscera. Similarly, 



166 Manual of Veterinary Microbiology. 

when it accidentally penetrates into the living tissues 
and finds there good conditions of vitality, it only 
multiplies locally and not in the blood, the oxygen 
of which destroys these microbes as soon as they en- 
ter this medium; on the contrary, some time after 
death it is found in the circulatory fluids. 

Large wounds, exposed to free contact with the 
air, are not easily contaminated on account of the 
anaerobic character of this bacillus. On the other 
hand, the latter readily implants itself in irregular 
contused wounds where the affected tissues are in 
way of necrosis. The following experiment of MM. 
Chauveau and Arloing is decisive in this regard. If, 
after the injection of a few drops of the virus into 
the jugular of a ram, the circulation is arrested in 
one testicle by bistournage, this testicle becomes the 
starting point of a fatal gangrenous process. 

When a wound becomes contaminated with the 
septic bacillus, if the local conditions are not adverse 
to its multiplication, an inflammatory process is seen 
to supervene the characters of which are those of the 
experimental oedema noted above. Absorption of 
the products elaborated by the bacilli, in other words, 
the septic intoxication, leads to general manifesta- 
tions of the disease which, sooner or later, terminates 
fatally. In fact, when 30 to 40 cub. cent, of the se- 
rosity of the oedema, deprived of its bacteria by fil- 
tration, are injected into the guinea pig, it leads to 
death in a few hours with symptoms of septicaemia. 

Transmission of the disease from one subject to 
another most frequently occurs by means of surgical 
instruments which have been contaminated by con- 
tact with an infected wound; this is the cause of the 



Microbic Diseases Individually Considered. 167 

epidemics which have been observed in mankind ; 
since the time that disinfectants came into general use 
this complication of wounds has become extremely 
rare. 

Septicaemias of the rabbit. 

The rabbit is very sensitive to the action of patho- 
genic microbes ; it succumbs in a comparatively short 
time to inoculation with the majority of these germs, 
and for this reason constitutes an important laboratory 
auxiliary. The experiments of Davaine, Coze and 
Feltz, and of Koch upon the virulence of putrid ma- 
terial were made principally with this rodent. Of 
these researches we have only to refer to those which 
relate to the experimental septicaemia of Koch ; the 
microbe peculiar to this disease shows, indeed, many 
characters in common with those of certain diseases 
of our animals, such as chicken cholera, duck cholera, 
pneumo-enteritis of the pig, and the epizootic of deer 
(wild-seuche). But, independent of all experimental 
conditions, rabbits may contract diseases of a similar 
nature, which prevail epizootically in their hutches 
and occasion serious losses. These, in contradis- 
tinction to the preceding, are designated spontaneous 
septicaemias. We will study here, in their essential 
points, first, the experimental septicaemia of Koch, and 
then the spontaneous septicaemias. The reader will 
readily notice the points of similarity of these differ- 
ent diseases. 

Experimental septicaemia of the rabbit. (Koch.) 
Koch produced this disease by the injection of a 
maceration of putrefied meat ; he obtained a putrid 
phlegmon, and death at the end of three days ; the 



168 Manual of Veterinary Microbiology. 

cedematous serosity at the periphery of the abscess, 
when inoculated in very small doses, transmits a 
septicaemia, fatal in twenty-four hours, to successive 
series of rabbits. 

The oadematous fluid and the blood contain, in 
large numbers, ovoid microbes 0-8// to I// in length, 
of which the extremities only take the color, so that, 
after staining, it affects the form of an 8. 

The disease is easily transmissible by inoculation to 
all species of birds ; according to Petri it may even 
prevail as an epizootic in chickens, ducks arid geese. 
It is not transmitted to the guinea pig. 

In the rabbit it gives rise to the following lesions : 
oadema at the point of inoculation, hemorrhagic 
patches on the peritoneum and in the lung, and en- 
largement of the spleen. Inoculated birds show a 
rapid emaciation with lowering of the body tem- 
perature; death, preceded by convulsions, arrives in 
less than twenty-four hours. The alterations con- 
sist in ecchymoses in the cellular tissue, abdominal 
effusion, petechise upon the intestine, infiltration of 
the lung, and the presence of a spumous mucus in 
the bronchi. The blood of birds transferred again to 
the rabbit in the smallest traces, reproduces the sep- 
ticaemia in the latter. 

Spontaneous septicaemias of the rabbit. 
Lucet has described a disease of these animals 
which prevailed in the hutches and occasioned serious 
losses. The subjects show inappetence, emaciation, 
torpor and temporary muscular spasms ; diarrhoea 
also occasionally supervenes, and death always quickly 
ensues. At the autopsy the blood is dark, the spleen 



Microbic Diseases Individually Considered. 169 

much enlarged and darkened ; the pleura and peri- 
toneum are the seat of exudative inflammation with 
fibrinous deposits, and there is slight abdominal 
effusion. 

The blood and organs contain a non-motile micro- 
coccus, isolated or associated in pairs, 0*7// to 0-9// in 
diameter; it takes the stain quite uniformly through*- 
out, but is not stained by the Gram method. It is at 
once aerobic and anaerobic, quickly loses its virulence 
in cultures left in contact with the air, and does not 
vegetate on gelatin or potato. 

The disease is transmissible from rabbit to rabbit, 
from the rabbit to the guinea pig and inversely, by 
inoculation, ingestion and by simple cohabitation. 
Its virulence becomes attenuated in the organism of 
the guinea pig, but regains its original strength on its 
return to the rabbit. In this last a culture which, 
from age, has lost a part of its virulence gives at first 
a local abscess. The disease studied by Lucet is not 
transmissible to the chicken. The food and dejec- 
tions are the vehicles of the germ and it is by their 
intermediation that the natural infection occurs. . 

Thoinot and Masselin have also studied a spon- 
taneous septicsemia of the rabbit which decimated 
the hutches at the Alfort school. The symptoms 
noted are loss of appetite and vigor, acceleration of 
respiration, and sometimes diarrhoea. The lesions 
consist in a dark color of the blood, deep wine color 
of the muscles, roseate or yellow effusion of the 
peritoneum and pleura, and albuminous urine. 

The disease was attributed to a micrococcus, single 
or associated in pairs, motile, presenting in birds the 
appearance of a short bacillus like the figure 8, of 



170 Manual of Veterinary Microbiology. 

which the two extremities, possessing more affinity 
for the coloring matter, are separated by a clear 
space. This microbe, once colored, is decolorized by 
the methods of Gram and of Weigert. It is found in 
large numbers in the blood and in most of the organs 
when the disease has lasted for some time; on the 
contrary, it is much rarer in very acute forms. 

Its culture is easy; it behaves as a facultative 
anaerobe and reproduces itself in the form of a mo- 
tile diplococcus recalling absolutely the microbe of 
chicken cholera. After twenty days exposure to the 
air, its cultures in bouillon have lost all virulence. 
When growing in bouillon the latter becomes turbid, 
then clear again ; gelatin is not fluidified and the cul- 
ture in this medium takes the form of a thick, white, 
glistening track, dentated at its margin. 

The disease is readily transmitted from rabbit to 
rabbit ; it is also inoculable to the guinea pig and to 
all species of birds, in which it occasions symptoms 
resembling those of avian cholera. When the virus 
is inoculated in the pectoral muscle it produces a se- 
questrum in all respects comparable to that caused 
by the inoculation of this last disease. 

Chicken cholera. (Fowl -cholera) 
This is an extremely fatal disease which prevails 
epizootically among farm -yard fowls. It is charac- 
terized by easily recognizable symptoms : the affected 
animals lose their accustomed vigor, generally as- 
sume the sitting posture, or move listlessly around; 
they are chilly and seek the sunshine, their plumage 
is ruffled, giving them the appearance of a ball of 
feathers; the comb becomes purple, bluish, or even 



Microbic Diseases Individually Considered. 171 

black. The appetite soon disappears ; an abundant 
diarrhoea ensues with expulsion of a glairy material 
and occasionally a viscid liquid may be rejected by 
the beak. Death supervenes in a few days, some- 
times preceded by convulsive movements; but the 
disease may be fulminating, and we then find fowls 
dead in their nests, the disease having produced its 
effects during the deposition of the egg. Cholera 
may also be of considerable duration and give rise to a 
slow emaciation of the affected animals ; in these cases, 
again, death is the usual termination of the disease. 

Fowl cholera is the cause of serious losses', hun- 
dreds of poultry sometimes dying in the same yard 
within a few weeks. The disease appears to have 
not always the same degree of malignity; this pe- 
culiarity is accounted for by the great mutability of 
the germ which occasions it. 

The changes found at the autopsy are quite con- 
stant: the blood is dark, usually tarry in appearance; 
however, it is not uncommon to find consistent clots in 
the cavities of the heart. The liver is large, dark, or 
sometimes of a rather light-brown color, and dotted 
with hemorrhagic patches; the intestines contain a 
quivering jelly-like mucus more or less adherent to 
the mucosa; the latter is inflamed, sometimes ulcer- 
ated, its alterations being more marked as the disease 
has been of longer duration ; in acute cases we es- 
pecially notice in this situation the presence of nu- 
merous petechise. The heart shows a characteristic 
lesion : its external surface is dotted with hemorrhagic 
points localized especially in the coronary groove ; the 
pericardial sac contains a quantity of fluid exudate 
and a gelatinous deposit adherent to the heart ; this 



172 Manual of Veterinary Microbiology. 

deposit is rarely absent. Miliary extravasations may 
also be found in the nerve centers. 

Fig 4 Microbe. The blood, the dejections 

and the pericardial exudate contain, 

u 

in large numbers, a micro-organism, 
very short and ovoid, resembling in 
shape the figure 8. It is a diplo-bac- 
terium, the extremities of which are 

less refringent and have more affinity 
1. Microbes of for the coloring matters than the mid- 
chicken cholera die part, which remains clear. It 

in figure 8 form; often ap p ears as a micrococcus ; this 
2. The same in . , L .. , . , -^ 

their real form 1S wne11 lts l n g axis corresponds with 
bacilli with clear the direction of the visual ray and the 
central space. (M. organism is seen on one of its ends. 
It is motile, its movements being very 
rapid in preparations of fresh blood.* It measures 
from 0-6// to 0-8/z in length by O3/* to 0-4/^ in thick- 
ness. It is a facultative aerobe, oxygen, however, be- 
ing favorable. to its multiplication. 

Action of physical and chemical agents. The bac- 
teria of chicken cholera are killed in fifteen minutes 
at 50, in ten minutes at 80, still more quickly at boil- 
ing temperature. By desiccation they are killed in 
a few days. Corrosive sublimate, at 1 to 5,000, kills 
them in one minute, carbolic acid, at three per cent, in 
six hours. They are not affected by the gastric juice. 
By the oxygen of the air they are first attenuated 
and then killed. 

Cultures. This germ grows well in bouillon in the 
presence of air; during its growth this medium 

* [Described in German works as non-motile. D.] 



Microbic Diseases Individually Considered. 173 

quickly becomes clouded and at the end of several 
days clears again by the deposition of the micro- 
organisms. 

Gelatin inoculated in lines becomes covered along 
the latter by a raised transparent pellicle ; inoculated 
by puncture it shows small gray colonies all along 
the needle track; it is not fluidified. 

The culture succeeds very indifferently on potato. 

Eesearch and coloration. The microbes of chicken 
cholera are easily distinguished, under strong mag- 
nification, in uncolored preparations of fresh blood; 
they appear as very refringent, mobile diplo-cocei. 
They must be stained with the hydro-alcoholic solu- 
tions as they are decolorized by the Gram method; 
Loffler's method is most suitable, especially when it 
is desired to study the microbes in sections. 

Experimental inoculations. The disease prevails 
spontaneously in and is successfully inoculated to all 
species of poultry: chickens, ducks, geese, pigeons, 
and turkeys; the pheasant and sparrow are also sus- 
ceptible. Subcutaneous inoculation and ingestion 
give results almost equally certain. The introduc- 
tion of the virulent matter into the pectoral muscle, 
by means of the Pravaz syringe, causes the formation 
of a sequestrum which is the more pronounced as 
the experimental disease is of longer duration. This 
sequestrum, however, is not characteristic of the dis- 
ease ; it may be seen after the injection of other germs 
(those of pneumo-enteritis of the pig, the spontane- 
ous rabbit septicsemia of Thoinot and Masselin, and 
a number of septic bacteria). 

The rabbit is extremely sensitive to chicken chol- 
era. The subcutaneous injection of a drop of blood 



174 Manual of Veterinary Microbiology. 

coming from a diseased chicken kills it in twenty- 
four hours. Hence, it is a delicate reagent for veri- 
fying the nature of an epizootic of fowl cholera, but, 
in order that this experiment should be of real value, 
it is evidently necessary to carefully avoid all causes 
of error which may result from the intervention of 
foreign germs. . The blood of the rabbit which suc- 
cumbs under such conditions is exceedingly rich in 
the specific germs. The rabbit can also be infected 
by way of the digestive canal. 

Inoculation of a drop of blood from a diseased 
chicken in the subcutaneous cellular tissue of the 
guinea pig causes an abscess which heals by evacua- 
tion of the pus; but inoculation into the blood results 
in death, the same as in the rabbit and chicken, by 
blood asphyxia. The pus of the abscess, in the 
guinea pig, is rich in microbes and its inoculation to 
chickens or to the rabbit reproduces the disease. 

It appears from a number of observations that the 
contact of virulent blood or of a culture with a 
wound can occasion in man, also, the formation of an 
abscess, and that the cat and the dog may consume, 
with impunity, chickens which have died from the 
disease. 

Etiology and pathogeny. The germ of chicken chol- 
era is deposited on the soil of poultry houses and 
yards with the fluid dejections of diseased fowls; 
their entrance into the organism of healthy subjects 
takes place by way of the digestive canal, the fowls 
taking up particles contaminated with the gerrns 
along with their food. The transmission of the dis- 
ease by ingestion has, moreover, been experimentally 
proved. Since desiccation and contact with the air 



Microbic Diseases Individually Considered. 175 

quickly kills the microbe, it is in moist poultry pens, 
in which the excretions accumulate, and in stagnant 
waters of the yards, that the virus is, for the most 
part, preserved. The germs of the disease may thus 
persist for a long time on one farm even after the re- 
moval of all diseased fowls. 

The microbe of chicken cholera, being aerobic, ex- 
tracts the oxygen from the blood and thus determines 
symptoms of asphyxia, which show themselves by 
the dark blue color of the comb, general coolness of 
the body, and hemorrhagic patches on the pericar- 
dium, liver, peritoneum, etc. By filtering a bouillon 
culture through plaster of Paris, M. Pasteur has ob- 
tained a liquid free from germs and which, inoculated 
to fowls, produces, temporarily, the most prominent 
symptoms of the disease, giving rise to a condition 
similar to that which succeeds to the absorption of a 
narcotic dose of opium. The animal is at first ex- 
cited, then its feathers become ruffled, the appetite is 
lost, it becomes somnolent, and, after several hours, 
emerges from its temporary stupor. Thus we see 
produced experimentally the intoxication which suc- 
ceeds to the multiplication of the specific bacteria in 
the blood. 

Attenuation Preventive inoculation. In cultivating 
the microbe of fowl cholera in the air and inoculating 
cultures of different ages, M. Pasteur observed that 
their virulence progressively diminished. Thus, by 
inoculating each time the same number of fowls with 
virus fifteen days old, one month, two months and 
over, he observed that the mortality induced by these 
different specimens of virus gradually decreased, and 
that, at a certain time, all the inoculated fowls sur- 



176 Manual of Veterinary Microbiology. 

vived. If at the time that a fowl is inoculated with 
a virus of a certain degree of virulence, this same 
virus is inoculated to culture media, it there repro- 
duces itself, communicating to its progeny the special 
pathogenic activity which it itself possessed; the at- 
tenuation of the germ is therefore, in this case, hered- 
itary. A series of cultures of progressively decreas- 
ing virulence can thus be prepared. 

The attenuation is connected with the action of the 
oxygen of the air; in order to preserve the virulence 
of cultures it suffices to exclude them from contact 
with the air; in contact with the latter they gener- 
ally become inoffensive at the end of two months. 

The attenuated virulence shows a series of degrees, 
from the normal virulence up to non-virulence. 
When the virus is enfeebled to such an extent that it 
no more kills chickens, it develops at the point of in- 
oculation in the pectoral muscle a local alteration, 
ending in the formation of a sequestrum which may 
either be eliminated or absorbed. Fowls, after recov- 
ery, are vaccinated against the later action of a more 
virulent or mortal virus. The immunity thus con- 
ferred is the more efficacious the more intense the 
vaccinal disease ; its duration appears not to exceed 
one year; the immunity from a first vaccination is 
strengthened by inoculation of a second more virulent 
vaccine.* 

The enfeebled virus can regain its virulence when 

* [Kitt has obtained immunity in chickens against the virus of 
chicken cholera by injection of the blood serum of previously im- 
munised chickens, as well as by injection of the albumen of the 
eggs coming from immune hens. (Centralbl. f. Bad. XIV, 25.) 
-D.] 



Microbic Diseases Individually Considered. 177 

it is passed in succession through small birds (cana- 
ries, etc.). 

Infectious enteritis of chickens. 

Under the name of infectious enteritis, Klein has 
described an epizootic disease of chickens which has 
much resemblance to cholera; the initial symptom is 
diarrhoea; the subjects are quiet but never show the 
somnolence so characteristic of cholera. Death oc- 
curs in twenty-four to thirty-six hours after the first 
manifestations of the disease. 

The intestinal contents, the blood and the splenic 
tissue contain a special bacillus measuring 0-8^ to 
1*6/* in length by O'B/j. to 0'4/j. in thickness. The 
chickens naturally become infected by ingestion 
of contaminated substances. Inoculation of the blood 
of a diseased chicken into the subcutaneous tissue of 
another, results in the death of the latter; but the 
inoculated animal remains well during the first five 
days and dies on the seventh to the ninth, whilst 
chickens similarly inoculated with fowl cholera suc- 
cumb in twenty-four to thirty-six hours. Finally, 
the bacillus of the disease described by Klein appears 
not to be pathogenic for the rabbit or pigeon, thus 
differing from that of cholera. 

Epizootic dysentery of chickens and ducks. 
M. Lucet, veterinarian at Courtenay, has studied 
another epizootic disease of domestic fowls, which he 
has named epizootic dysentery. of chickens and ducks; a 
summer disease as deadly as the two preceding, this 
dysentery especially attacks young chickens of the 
same year, and gives rise to symptoms which strongly 
recall those of cholera : somnolence, lassitude, inap- 



178 Manual of Veterinary Microbiology. 

petence, diarrhoea, chilliness, ruffling of the plumage, 
etc. The temperature, at first high, afterward de- 
scends one to two degrees below the normal figure ; 
the animal dies on the ninth to the thirteenth day after 
the beginning of the disease, occasionally much later. 

The bacteria, which are present in most of the 
lesions and especially in the intestine, show them- 
selves in the form of short bacilli 1-2^ to 1*8^ "in 
length, occasionally isolated, more frequently united 
in pairs, motile, at once aerobic and anaerobic, rap- 
idly becoming attenuated in cultures. 

The disease is communicated, by ingestion of viru- 
lent products, from chicken to duck, and inversely. 
Inoculation of cultures also causes it in the same fowls ; 
but ingestion of cultures only produces the disease 
when the diet is changed at the same time. The 
pigeon and the guinea pig are refractory ; the rabbit 
takes the disease only by intravenous inoculation ; 
subcutaneous injection remains without effect. 

Duck cholera. 

Ducks are liable to contract chicken cholera ; in ad- 
dition, they may be attacked by another contagious 
disease which is also characterized by diarrhoea, 
emaciation of the affected subjects, and by its usually 
fatal termination ; it has been described under the 
name of duck cholera. 

This disease is caused by the multiplication in the 
blood of a bacterium presenting the greatest mor- 
phological analogies with that of chicken cholera; it 
is oblong, short, and appears bi-lobed in stained 
preparations on account of the greater affinity of its 
extremities for the coloring matters. The microbes 



Microbic Diseases Individually Considered. 179 

of duck cholera are a little larger than those of 
chicken cholera. Like the latter they do not admit 
of double staining. 

Their culture on the different solid media is richer 
than that of chicken cholera. This peculiarity is es- 
pecially noticeable upon potato, which is a very bad 
field for the growth of the organism of chicken 
cholera, whilst that of duck cholera grows very well 
on this medium. 

"Whilst chicken cholera is pathogenic for ducks, 
duck cholera is inoffensive for chickens and pigeons. 
The latter disease does not even kill all ducks with 
the same rapidity ; some species resist for a longer 
time than others. 

The rabbit succumbs to duck cholera as well as to 
chicken cholera, but requires a larger dose of the 
former than of the latter. 

It might be thought that the virus of duck cholera 
was an attenuated form of that of chicken cholera. 
But inoculation of the former to the chicken ought 
then to vaccinate against the latter disease, which is 
not the case. 

The penetration of the germs, in the case of spon- 
taneous contamination, takes place by the digestive 
canal ; this mode of infection has also been demon- 
strated experimentally. The experimental disease 
can also be communicated by hypodermic injection.* 

* [Several other microbic diseases of fowls have been described : 
Vibrio-cholera of chickens ( Vibrio Metschnikovi, Gameleia) in 
Russia, caused by an organism presenting morphological and 
cultural resemblances with that of Asiatic cholera ; septicaemia 
in geese (Spirochsste anserina, Sakharoff) in swampy regions of 
Transcaucasia a spiral microbe resembling that of relapsing fever 



180 Manual of Veterinary Microbiology. 

Bacteridian charbon. 

This is an infectious and contagious disease caused 
by the bacteridium.* 

The disease shows itself by a profound adynamic 
fever, with more or less marked stupor of the affected 
animals. The blood is much changed, viscid, and the 
plasma, loaded with the coloring matter of the cor- 
puscles, communicates to the mucous membranes a 
dull yellow tint ; sometimes visible hemorrrhages oc- 
cur: nasal and conjunctival petechiae, bleeding from 
the lungs and bowels, hematuria. The intesti- 
nal lesions in the horse "often give rise to more or less 
violent symptoms of colic, and this complication, 
considered too exclusively, frequently interferes with 
the diagnosis of the essential disease. 

At the autopsy the blood is found to be deoxygen- 
ated, viscid, incoagulated, the corpuscles are altered, 
agglutinated, and the plasma colored red. The inter- 
nal tunic of the bloodvessels and of the heart is also 
often stained red; petechise are found on the heart, 
lung, pleura, and peritoneum. The spleen is much 
enlarged ; its borders, clear cut in the normal condi- 
tion, have become rounded ; its surface is often lumpy, 
its consistence soft and friable, and its meshes infil- 
trated with extravasated blood. The intestines are 

of man is found in the blood, not cultivated ; epizootic disease 
of grouse (Klein) in England ; epizootic pneumo-pericarditis in 
turkeys (McFadyean) in England. D.J 

* [The term " Bacteridium " or " la bacteridie," derived from the 
genus Bacteridium of Davaine's early classification is retained by 
French writers as a specific name for the bacillus anthracis. " Bac- 
teridian charbon " or " charbon," refers to the disease produced 
by this organism. Synonyms; anthrax, splenic apoplexy, etc. 
Ger. Milzbrand. D.J 



Microbic Diseases Individually Considered. 181 

sometimes the seat of intense congestive and hemor- 
rbagic lesions, and in some cases the lymphatic glands 
of the different regions are in the same condition, and 
also enlarged to twice or three times their normal 
size. Similar lesions may also be found in the kid- 
neys, raeninges, etc. 

The species which are liable to contract the spon- 
taneous disease are the sheep, goat, ox, and horse; this 
last less readily becomes infected than the first; it can 
consume with impunity foods which occasion the dis- 
ease in the others. Charbon is also met with in the 
carnivora of menageries (lion) when they are fed with 
the flesh of animals which have died of this disease. 
Exceptionally the dog and the pig become infected in 
the same way. Algerian sheep are refractory even 
when they are born in other countries. 

Man is unfortunately also liable to contract this 
disease; the latter then receives different names ac- 
cording to the mode of penetration of the bacteridium 
and the initial lesion which it determines : 

1st. Malignant pustule, the most frequent form, con- 
secutive to the accidental insertion of the virus into a 
cutaneous wound, develops in workmen who cut up, 
dress, or retail charbonous meat. 

2d. Pulmonary charbon, the rarest form, develops in 
consequence of the inhalation of dust charged with 
the bacteridia, or rather with their spores, in work- 
men who handle wool or skins coming from the bod- 
ies of charbonous subjects: 

3d. Intestinal charbon, consecutive to the consump- 
tion of charbonous meat. 

Characters of the bacteridium. The microbes of 
charbon are straight, cylindrical rods; they are iso- 



182 Manual of Veterinary Microbiology. 

lated or associated in twos or threes, rarely more, 
the limits of the articles being then marked by one or 
more articulations or clear zones distinctly travers- 
ing the polybacillar filament; often two contiguous 
elements have commenced to detach themselves and 
form between them a large open angle with the articu- 
lation partly disconnected, forming its apex. 

The different segments of the same filament are of 
equal length ; each segment is very slightly swollen 
at its extremities. Their dimensions vary from 5/z to 
10// in length by I/* to 1*5// in thickness. The com- 
pound filaments are never of great length in the 
blood of animals dead of charbon on account of their 
constant collision with the blood corpuscles. In arti- 
ficial cultures, on the contrary, they become consider- 
ably elongated, important changes at the same time 
being seen to take place in their substance; their 
Fig. 5. homogeneous contents become 

I. A modified, condensing in the form 
of spores ; the latter are ovoid cor- 
puscles, highly refringent, less thick 
than the filament itself and conse- 
quently never produce bulgings in 
the latter. The formation of the 
spores is followed more or less 
' " quickly by their liberation, through 
4. The same in a cul- tne disintegration of the filaments, 
ture, sporulated. (M. Reproduction of the bacteridium, 
and L> ) therefore, takes place only by fis- 

sion in the bodies of diseased animals during life; in 
cultures the transversal division, only brought into 
view by staining, is followed by sporulation, but only 
between certain limits of temperature ; it begins above 




Microbic Diseases Individually Considered. 183 

12 and ceases at 40, and is the more active in pro- 
portion as the nutritive fluid becomes more exhausted. 

The bacillus of charbon is non-motile and aerobic ; 
the presence of oxygen is, indeed, absolutely neces- 
sary for sporulation. 

Action of physical and chemical agents. A tempera- 
ture of 45 arrests the vegetation of the charbon 
bacilli; at 42-5 they multiply only by fission, no 
longer giving birth to spores, and they also become 
attenuated. 

The rods are killed by several minutes exposure to 
a temperature of 50 ; the spores, however, are not 
influenced by this temperature ; in the moist con- 
dition they resist 80 and when dry support with im- 
punity a temperature of 100. 

The bacteridia, in the condition of mycelium, that 
is, the rods, are destroyed by putrefaction ; hence 
they quickly disappear from carcasses abandoned to 
the air. But this is not the case for those in which 
the viscera have been immediately removed, and more 
especially for bodies of charbonous animals which 
have been bled and dressed. Putrefaction, in this par- 
ticular case, is much more tardy, and it is possible 
to find the charbon bacillus in the blood several days 
after death. This is a fact of very great practical 
importance and one which meat inspectors ought 
carefully to notice. 

It would seem that, the bacilli of charbon being 
destroyed by the putrefaction of the cadavers, the 
latter would not constitute a source of danger for 
later contaminations. But after the death of the dis- 
eased animals all the bacilli are not destroyed ; those 
which come into contact with the external air form 



184 Manual of Veterinary Microbiology. 

spores and these are completely resistant to decom- 
position. In bodies from which the skin has been re- 
moved the exposure of a large number of bacilli in- 
volves a more abundant production of spores than in 
those which have riot been skinned. 

The bacilli accidentally deposited upon the soil by 
diseased animals, finding themselves in contact with 
the air and at a suitable temperature will also form 
spores, which are preserved for a long time on the 
surface of the vegetation. According to some au- 
thors, these spores themselves can pass through the 
different phases of their evolution in the soil and 
give rise to new generations. 

The bacteridia retain their virulence for a length- 
ened period in the blood when this liquid has been care- 
fully collected and protected from the invasion of the 
germs of decomposition. Dried blood also long re- 
tains its virulence; the bacilli are brought to the con- 
dition of latent life and will again multiply when un- 
der good conditions of humidity and temperature. 

Antiseptics act very differently on the bacilli and 
on their spores. Thus one-fourth to one-half per 
cent solution of carbolic acid kills the rods, whilst a 
five per cent solution is required to kill the spores. 
The bacteridia are killed by carbonic acid, by com- 
pressed oxygen, and by absolute alcohol, whilst the 
spores resist these agents. 

Cultures. The bacilli of charbon readily multiply 
in artificial media at temperatures between 12 and 
43, and in contact with the air. The most favorable 
temperature is about 38. 

In bouillon, after the first day, white mucoid flakes 
are seen suspended in the fluid ; these do not become 



Microbic Diseases Individually Considered. 185 

dissociated by shaking. They slowly increase in size, 
still remaining united; they are formed of filaments 
of great length intwined with each other like the 
threads of cotton wadding. After a certain time 
these filaments produce spores and break up ; the lib- 
erated spores fall to the bottom of the vessel where 
they look like fine sand. 

Inoculated to gelatin by puncture, the charbon bacillus 
forms a culture no less characteristic; along the deep 
track a white line appears from which project hori- 
zontally ramifying branches, so that the whole simu- 
lates the branch of a tree with its divisions and sub- 
divisions; on the surface a white layer is formed 
which slowly fluidifies the gelatin and gradually in- 
creases in thickness. 

The colonies which develop on gelatin plates are 
formed of tufts of interlacing filaments showing ar- 
borescent prolongations at their periphery. 

On agar the bacteridium grows as upon gelatin.* 

On potato it produces a dry crust of a white color. 

Virulent cultures contain various toxic substances: 
a toxalbuminoid, precipitated by alcohol (Nankin, 
Brieger, Fraenkel), and an alkaloid (Martin). 

Research and coloration. ^The charbon bacillus is 
found in the blood of animals which have died from 
the disease, and in the local cedema consecutive to ac- 
cidental or experimental inoculation. It is easily 
recognized in fresh unstained blood preparations in 
the form of very clear articulated or non-articulated 
rods, lying motionless between the corpuscles. It 

* [But the growth in agar tubes is ID no way characteristic. D.] 
16 



186 Manual of Veterinary Microbiology. 



Fig. 6. 



readily takes the different aniline stains; the blood 
may be stained with aqueous solutions either directly 

or after being dried 
on the cover glass; 
in this last case the 
bacteridia are gene- 
rally shorter and 
more slender. The 
double stains of 
Gram and Weigert 
give excellent re- 
sults.* 

Examination of 
fresh blood is espe- 
cially to be recom- 
mended here, as it 
enables us to estab- 
lish the immobility 
of the bacilli and the 
alterations of. the blood corpuscles. In examining 
the blood of a carcass for bacteridia the specimen 
must be taken from a deep vein some distance from 
the peritoneum, because, in this disease, the invasion 
of microbes coming from the abdominal viscera is 
more rapid as the charbon bacillus, being aerobic, has 
deprived the blood of its oxygen. 

Experimental inoculations. Inoculations may be 
made by subcutaneous injection, by intra- vascular 

* [The Gram stain is especially applicable to the staining of sec- 
tions. On cover-glass preparations the characteristic appearance 
of the bacillus is best brought out by the use of aqueous or hydro- 
alcoholic solutions. Loffler's stain, followed or preceded by eosin, 
gives excellent results. D.] 




Anthrax bacilli in the blood. 
(Cover-glass preparation) . Kitt. 



MicroUc Diseases Individually Considered. 187 

injection, or, more simply, by superficial scarifications 
of the integument. The disease can also be artifi- 
cially communicated by ingestion, or even by inhala- 
tion. To confirm a diagnosis hypodermic inoculation 
and scarifications are amply sufficient. 

The substances to be inoculated are represented by 
the blood or its serum and the diluted pulp of organs 
rich in vessels : spleen, liver, lymphatic glands, etc.; 
in laboratories cultures are also used for inoculation. 

The disease is transmissible by inoculation to nearly 
all the domesticated animals. Mice, guinea pigs, and 
rabbits are the animals which are most frequently 
employed for experimental inoculation for diagnostic 
purposes. The white rat possesses an almost com- 
plete immunity, although under certain circumstances 
it can also be succesfully inoculated. 

The dog takes the disease only when large doses 
are employed or when the virulent substance is intro- 
duced into the vascular system. Fowls are refractory 
to both the natural and experimental disease ; but this 
immunity is easily overcome : cooling a fowl to 38, 
by placing it in a current of cold water, is sufficient 
to induce the development of the disease after inocu- 
lution. These fowls, when again warmed, recover. 
The temperature of fowls is, therefore, naturally too 
high to admit of the pullulation of the charbon bacil- 
lus. On the contrary, that of frogs is too low, and we 
only succeed in infecting these animals with charbon 
when they are heated in a bath to 35. 

In the rabbit and the guinea pig the inoculation is fol- 
lowed, at the end of ten to fifteen hours, by a marked 
local osdema ; the temperature is elevated from one to 



188 Manual of Veterinary Microbiology. 

two degrees. Nevertheless, the appetite only fails a 
few hours before death. The latter supervenes in 
thirty-six to forty hours in the guinea pig, forty-eight 
to sixty in the rabbit ; the animal dies in coma or 
after slight convulsions, and always with a marked 
depression of temperature. 

The autopsy of animals which have succumbed to 
the inoculated disease is interesting : the inoculated 
place is the seat of a jelly-like cedematous infiltration, 
of a faint red color. This oedema is found in natural 
cases of charbon when the penetration of the bacte- 
ridia has occurred at an erosion of the digestive mu- 
cous membrane : gloss-anthrax, charbonous angina. 
The corresponding lymphatic glands are tumefied, ec- 
chymosed, and surrounded by an cedematous zone. 
The osdematous fluid and the blood are charged with 
bacteridia. The spleen is much enlarged, diffluent, 
and dark in color.* 

* [We have little reliable information as to the extent to which 
anthrax prevails in the United States and Canada. Local out- 
breaks have been reported from many different quarters, but the 
microscopical and laboratory investigations necessary to the iden- 
tification of the disease have generally been lacking. A disease 
known to the natives as "charbon" prevails in some yeans as an 
epidemic among mules in certain parts of Mississippi (also re- 
ported in Louisiana and Arkansas). This disease was investigated 
and described as charbon by Veterinarian Creelman, of the Mis- 
sissippi Agricultural Experiment Station, and the diagnosis con- 
firmed by the laboratory investigations of Dr. Connoway, of the 
Missouri Experiment Station. (Miss. Agric. Exper. Sta. Bull. 
JVb.ll.) 

Material from the same source (obtained from a cow), examined 
at Washington, was found to contain the anthrax bacillus. (An. 
Rep. Bureau of Animal Industry, 1889-1890, p. 43.) 

An extensive outbreak of anthrax, in Illinois, is reported by 



Microbic Diseases Individually Considered. 189 

Etiology and pathogeny. Diseased animals dissem- 
inate the virus in their faecal dejections and urine, 
which are often mixed with blood and therefore 
with bacteridia ; frequently, also, the blood escapes to 
the exterior directly by the nasal cavities. Their car- 
casses, all parts of which contain the virus, are more 
important sources of contagion ; hence, when these 
carcasses are buried without any precautions being 
taken to destroy the charbonous germs, they may 
become the starting point of fresh cases. These 
bodies become more directly dangerous when the 
flesh is used for food. 

Contagion by direct contact is comparatively rare ; 
it occurs when a person inoculates himself with char- 
bon at a cut or any other recent wound, in cutting up 
a charbonous carcase; it also occurs when the dis- 
ease follows the consumption of diseased meat, and 
finally, by transmission, now well established, from 
the mother to the foetus. 

Contagion most frequently takes place by indirect 
contact, the germs having been previously distributed 
in the surrounding media. The floor of sheds, the 
soil and vegetation of fields occupied by diseased ani- 
mals, become soiled by their dejections; the germs 
thus deposited may contaminate the litter and fodder 
and thus gain entrance to the alimentary canal of 
herbivora. It is not necessary in this case, as was at 
first believed, that the animals ingest at the same time 

Dr. Trumbower. (Rep. Board of Live Stock Commissioners of Ills., 
1893.) 

A few cases in cattle, in which the diagnosis was confirmed by 
competent bacteriologists are reported from Delaware. (Fifth An. 
Rep. of the Del. College Agric. Experiment Sta., 1892, p. 45.) D.] 



190 Manual of Veterinary Microbiology. 

substances capable of wounding the mucosa, thus 
opening the way for the bacteridia. It has been 
shown that the spores of charbon can be absorbed in 
the absence of all intestinal wounds. However, the 
non-sporulated bacilli are killed by the gastric juice 
so that their action can only take effect in the passages 
anterior to the stomach, and by a solution of contin- 
uity. Experiment has shown that the addition to 
contaminated fodder of substances capable of ex- 
coriating the mucosa (thistles, husks of barley) in- 
creases the mortality from charbon. At the wounds 
thus produced there is first developed a tumor, from 
which invasion proceeds by way of the lymphatics. 

"We have seen that, if the charbon bacilli have them- 
selves great resistance, this faculty is possessed to a 
still greater degree by their spores. It is, morever, 
under this last form that the contagion of charbon 
persists on certain farms in such a way as to give the 
disease an endemic character. The contagion having 
once been deposited on a field, even on a limited part 
of the same, it remains there for years, contaminating 
the vegetation which grows upon it. The spores have 
been found on the surface of the ground above a grave 
closed for twelve years. The disease occurs during 
the pasturing season or in winter, according as the 
herbage is consumed while growing or after harvest- 
ing, in the form of hay. The most frequent cause of 
these endemic foci resides in the manner of burial of 
charborious carcases : in spite of the layer, of greater 
or less thickness, by which they are covered, the spores 
produced in these bodies * find their way to the sur- 

* [It seems to be well established that spores are not formed in 



Microbic Diseases Individually Considered. 191 

face. It is asserted that this migration is chiefly due 
to the agency of earth-worms. We know that these 
worms, in excavating the passages through which they 
wander, swallow a certain quantity of earth which 
they then expel in the form of tortuous rolls ; we also 
know that they make regular nocturnal peregrina- 
tions to the surface of the soil in order to feed on the 
herbage. Now, M. Pasteur has found, in these little 
rolls of earth deposited above a grave containing a 
charbonous carcase, the spores or germ corpuscles 
of the disease. These spores do not necessarily always 
remain limited to the part of the field on which they 
have been deposited ; they may be more or less widely 
disseminated by winds, and more especially by water. 
If we admit as established the possibility of the 
multiplication of the bacilli of charbon in swampy 
soil we can readily account for the persistence of the 
disease in certain localities. 

The charbon bacilli may penetrate into the blood 
directly or indirectly ; in this last case they first make 
their way into the tissues, multiply there, and excite 
an inflammatory engorgement, then gain the lym- 
phatic glands and, finally, the blood. They then act 
by a complex mechanism. By reason of their aerobic 
character they rob the blood of part of its oxygen and 
thus give rise to asphyxia. This asphyxiating action, 
however, is contradicted by the investigations of 
M. Chauveauwho has demonstrated the presence of a 
normal proportion of oxygen in the blood of a sheep 

the blood or organs of buried cadavers (Feser, Kitasato, Kitt) ; they 
are found, however, on the surface of these bodies where they are 
soiled with blood, excretions, etc., and also in the digestive canal 
both before and after death. D.] 



192 Manual of Veterinary Microbiology. 

in the last stage of charbon. It is especially by its 
secretions that the bacteridium acts upon the organ- 
ism. The nature of these substances is as yet im- 
perfectly understood, but their action is demonstrated 
by the following experiment: M.Cbauveau transfused 
to a healthy sheep a large quantity of blood coming 
from a sheep affected with charbon, and, as a conse- 
quence, noticed the development of the general symp- 
toms of the disease. This immediate result can only 
be explained as a chemical poisoning, an opinion 
which was, moreover, confirmed by the microscopic 
examination, which showed the disappearance of the 
bacilli from the transfused sheep. It is to this in- 
toxication that we must ascribe the phenomena of 
nervous depression or temporary excitement, which 
are so marked in charbonous subjects. 

Cbarbonous blood filtered through porcelain acts on 
the red corpuscles of normal blood, rendering them 
viscid and glutinous. This property is communicated 
to it by the substances secreted by the charbon bacilli 
and it explains the peculiar alteration of the blood 
in this disease. 

Finally, the bacilli act mechanically ; on account of 
their number and the abnormal viscidity of the blood, 
they form plugs in the interior of the capillary ves- 
sels, thus occasioning blood stases and superficial and 
deep hemorrhages. This is undoubtedly the cause of 
the final passage of the bacteridia into the milk, 
urine, and through the placenta ; it is also the cause 
of the large and characteristic swelling of the spleen, 
as well as of the hemorrhages and general circulatory 
disturbance. 

Attenuation. Vaccinations. The virulence of char- 



Microbic Diseases Individually Considered. 193 

bon cultures is very stable ; this is due to the pres- 
ence of the spores which are little subject to change. 
When it is desired to attenuate these cultures it is 
necessary to begin by preventing the formation of 
spores. Pasteur, succeeded in this by cultivating the 
charbon bacilli at the temperature of 42 to 43. 
Multiplication of the bacilli still continues, but spores 
are no longer formed. Now, if such cultures are 
kept in contact with the air their virulence rap- 
idly diminishes ; after twelve days they no more kill 
adult guinea pigs, and vaccinate the rabbit and the 
sheep ; the power of vegetation, however, still per- 
sists, and is absolutely extinguished only at the end .of 
one and a half months, on an average. The bac- 
teridium which has become asporogenous at the tem- 
perature of 42 to 43 then loses its virulent properties, 
retaining only those of an ordinary saprogenic mi- 
crobe, and, finally, it loses all vitality. 

The culture loses its pathogenic power little by lit- 
tle ; it ceases to be fatal first for the large animals, 
then for small adults, and finally for small animals 
only a few days old. Now, each degree of virulence 
can be perpetuated separately by cultivating at 42 
to 43 the different varieties obtained, each of these 
varieties transmitting its special virulence to its de- 
scendants. The degree of attenuation of each of 
these varieties can be preserved if the precaution be 
taken to frequently transfer to fresh culture media ; 
but it is not possible to entirely avoid the attenuation 
which finally takes place. In order to definitely fix 
these varieties it suffices to return them to 37 ; they 
then form spores possessing, in embryo, the special 
17 



194 Manual of Veterinary Microbiology. 

virulence of the bacilli from which they came and 
capable of transmitting this virulence to new gen- 
erations of bacilli cultivated at 37. 

The least virulent varieties obtained produce a be- 
nignant disease which leaves behind it immunity for 
the varieties less attenuated, and thus are obtained 
vaccines of different degrees of intensity, which may 
be employed in succession. In practice two 'vaccines 
only are used, of which one or two drops are in- 
jected at an interval of ten days. 

In the general part we have described the other 
methods of attenuation of the charbon bacillus by 
compressed oxygen, by heat and by antiseptics and 
the vaccination processes which have been derived 
from them. 

Symptomatic charbon* 

This disease, formerly confounded with charbon 
properly so called, has been separated from the latter 
as a result of the investigations of MM. Arloing, 
Cornevin, arid Thomas. It is characterized, first, by 
the symptoms of a more or less intense fever and by 
the appearance of a specific tumor upon the body, 
neck, or upper part of the limbs. This tumor is al- 
most constantly found in the muscular masses ; it con- 
sists, at first, of a painful and progressive inflamma- 
tory engorgement, of firm and uniform consistence ; 
it rapidly extends in area and in depth and, later, be- 
comes insensible, crepitant, and resonant at its center 
(emphysemato-gangrenous tumor). The general symp- 

* [Also occasionally referred to in this work as " bacterial char- 
bon." Synonyms : Symptomatic anthrax, black-quarter, infectious 
emphysema; iaf . Sarcophysema haemostatic umbovis; Ger. Rauseh- 
brand. D.] 



Microbic Diseases Individually Considered. 195 

toms become aggravated with the progressive evolu- 
tion of the local lesion and the subject succumbs in 
thirty-six to forty-eight hours. When the disease has 
been somewhat prolonged, other tumors with the 
same characters not infrequently supervene. The dis- 
tribution of these secondary tumors is not in relation 
with that of the lymphatics ; their development ap- 
pears to take place through the intermediation of the 
blood. 

At the autopsy the local lesion is the predominant 
feature, the invaded muscles are friable and more or 
less darkened in color (charbon) ; their fibrous bundles 
are readily dissociated ; the fibers retain their striation 
but their contents is broken up into hyaline, vitreous 
blocks (hyaline degeneration). The intra-muscular 
connective tissue, as well as that which surrounds the 
muscular masses, is thickened and infiltrated with a 
yellowish serosity ; this oedema sometimes assumes con- 
siderable proportions. The formation of gas, inherent 
to the life of the germ, causes the detachment of the 
tissues, a true localized emphysema in the central part 
of the tumor. The gases produced are, chiefly, car- 
bonic acid and marsh gas. 

The lymphatic glands in relation to the tumor are 
reddened, ecchymosed and infiltrated. 

After death the bodies very rapidly putrefy. 

The animals which spontaneously contract the dis- 
ease are cattle, sheep and goats. The receptivity of 
the first is not the same at all ages ; calves of less than 
six months do not contract the natural disease, and 
cattle of over four or five years also seem to escape. 

Microbe. The pathogenic agents in this disease are 
straight rods, isolated or occasionally associated in pairs, 



196 Manual of Veterinary Microbiology. 

measuring in the adult, non-sporulated condition 5/* 
to Sju by 1/j, ; after fruiting they may attain larger di- 
mensions IQfj. by 1-3/z; as long as their contents are 
homogeneous they are cylindrical, but their form 
changes with the appearance of the spores ; generally 
the spore is terminal and gives the bacillus a bell- 
clapper or club-shaped appearance; sometimes it is 
central, the bacillus then becoming spindle-shaped. 
The spore is solitary, ovoid, and very distinct; it oc- 
cupies one-third of the length of the element. 

The bacillus Chauvoei is endowed with oscillatory 
motion. It is strictly anaerobic; hence we should 
not expect to find it in the blood, at least during life ; 
after death, when the oxygen is no more renewed by 
the pulmonary exchanges, it penetrates into this fluid. 
Action of physical and chemical agents. 
The virus withstands extreme cold and 
^^A | after being dried also resists for a con- 
^* siderable time the action of high tern- 
Bacilli of symp- P eratures ; ^ is not destroyed by ex- 
tomatic charbon, posure for two hours to 80, or for 
non-sporulated twenty minutes to 100; on the other 

and sporulated. hand, it perishes in two minutes in 
(M. and L.) -, .-.. 

boiling water. 

The serosity dried at about 35 retains its virulence 
for more than two years. 

Putrefaction has no effect on the bacillus of symp- 
tomatic charbon. 

The bacilli, when they have escaped from the ca- 
davers and been deposited upon the soil, preserve 
their virulence* for a long time if the external condi- 
tions permit of their rapid desiccation ; under other 
conditions they are more or less quickly attenuated and 



Microbic Diseases Individually Considered. 197 

finally destroyed by the oxygen of the air. The same 
thing occurs when they have penetrated to a slight 
depth in permeable soil. But when they are buried 
at a sufficient depth, especially in a compact clayey 
soil, they are preserved for a considerable time on ac- 
count of the absence of oxygen and light, and they 
may even, by virtue of their anaerobic faculty, propa- 
gate themselves there. 

Strong antiseptics are fatal to the bacillus Chauvoai ; 
Arloing, Cornevin, and Thomas have observed this 
toxic action after twenty-four hours' contact with sub- 
limate, at 1 to 5,000, chlorine gas, and two per cent 
solution of carbolic acid ; on the other hand, sulfurous 
acid, quick lime, and 90 per cent alcohol do not de- 
stroy its virulence. 

Cultures. Cultures of this microbe are very difficult 
to obtain ; they are only possible when excluded from 
oxygen, in a vacuum, or in presence of an inert gas, 
such as carbonic acid.* Bouillons to which have 
been added glycerin and ferrous sulfate, or gelatin 
and sugar, are the media to be preferred. The fluid 
rapidly becomes turbid and the seat of an intense dis- 
engagement of gas ; it exhales a pronounced odor of 
rancid butter. 

Attempts at cultivation on solid media have also 
been successful. Upon gelatin there are produced 
spherical verrucose colonies, which fluidify the me- 
dium and give rise to a lively production of gas ; cult- 
ures upon agar have a penetrating acid odor. Viru- 
lence does not long persist in generations obtained by 

* [" Grows in an atmosphere of hydrogen but not in carbon di- 
oxide." Sternberg: Manual of Bacteriology, p. 494. D.] 



198 Manual of Veterinary Microbiology. 

artificial culture ; it becomes obliterated in four or five 
transfers. 

The material for inoculation of culture media may be 
taken from the juice obtained by scraping the center 
of a tumor, from the peritoneal serosity, and from the 
blood ; but they only appear in this last fluid after 
death ; as they are present in small numbers it is well 
to allow them to multiply by keeping a quantity of 
this blood in the incubator for twenty-four hours. 

Research and coloration. The bacillus Chauvoei ex- 
ists in great abundance in the muscular tumor, sus- 
pended in the fluid with which it is infiltrated and 
which is interposed between the contractile elements. 
In the last moments of life and after death it is also met 
with in small numbers in the blood ; finally, it exists in 
abundance in the bile and in the peritoneal serosity. 

Simple methods of staining are alone successful; 
the various hydro-alcoholic solutions are available for 
this purpose, but Loffler's method should be preferred 
for sections. . 

Experimental inoculations. The species to which it 
is possible to communicate the experimental disease 
are the ox, sheep, goat, and guinea pig. The rabbit is 
refractory. The ass and the horse only contract a local 
engorgement. 

The receptivity of the various species for this bacillus 
sufficiently differentiates it from that of Pasteur's sep- 
ticsemia which it much resembles in its physical char- 
acters, staining proclivities, and its anaerobic faculty, 
and in the emphysemato-gangrenous lesions which it 
occasions. The septic bacillus is pathogenic for all 
species except the ox ; the bacillus of symptomatic 



Microbic Diseases Individually Considered. 199 

charbon is pathogenic only for ruminants and the 
guinea pig. 

Dermic inoculation with the lancet or by superficial 
scarifications is nearly always unsuccessful, whilst the 
introduction of the virus into the subcutaneous cellu- 
lar tissue or into the muscular tissue gives positive re- 
sults. However, it is necessary to take into account 
the dose injected and the place in which the injection 
is made. Yery small doses do not produce the dis- 
ease but confer immunity; similarly, a dose which 
would be fatal if injected in a favorable place is inof- 
fensive when inoculated in the cellular tissue of the 
tail and of the extremity of the limbs. The experi- 
ments of M. Arloing have shown that this local im- 
munity, which had been established by clinical obser- 
vation, depends upon the greater density of the con- 
nective tissue and on the lower temperature of these 
regions. He succeeded in overcoming this immunity 
by heating the region or by lacerating its cellular 
tissue. 

After the insertion of the virus in the subdermic tis- 
sue there results a painful and progressive inflamma- 
tory engorgement which extends to neighboring re- 
gions and in the guinea pig leads to death in twenty- 
four to forty-eight hours. 

The intra-venous inoculation of small doses, but 
doses which would be fatal by the subcutaneous 
method (three to five drops of juice in young bovines, 
three-tenths of a drop in sheep), excites a rather in- 
tense febrile reaction ; tumors are not produced unless 
some of the virus has been deposited in the cellular 
tissue surrounding the bloodvessel. The subject thus 
inoculated is vaccinated against the temporary reac- 



200 Manual of Veterinary Microbiology. 

tion of a second intra-venous injection, and also against 
the subcutaneous inoculation of an otherwise mortal 
virus; the reaction of the latter then consists only in 
the formation of a curable abscess, the pus of which 
contains the virulent germs. 

If the dose which can be tolerated in the vessels is 
exceeded, the typical disease ensues with the develop- 
ment of tumors. Similarly, if, after the intra- vascular 
injection of a vaccinating dose, a hemorrhage is pro- 
duced in the connective tissue, a specific, fatal tumor 
appears in the place of the solution of vascular con- 
tinuity, the bacilli having penetrated into their media 
of predilection. 

The disease is transmissible by way of the respira- 
tory passages, the result being the same as by the blood- 
vessels. Finally, the disease can be transmitted by 
way of the intact digestive canal if the virus is very 
active ; the tumors then appear in places remote from 
the point of entry. 

Etiology and pathogeny. Symptomatic charbon is 
endemic in certain countries; it prevails especially 
during summer, its ravages being however less im- 
portant than those of bacteridian charbon. Animals 
inoculate themselves accidentally, and as small doses 
confer immunity and this is transmitted from the 
mother to the foetus, it results that part of the animals 
exposed to the contagion escape its fatal effects. But 
those which at first receive a sufficient dose quickly 
succumb. 

The virus seems to be capable of entering by differ- 
ent ways. Wounds of the external integument in 
favorable regions are more especially suited to its evo- 
lution ; but the inhalation of dust charged with dried 



Microbic Diseases Individually Considered. 201 

virus, as well as the ingestion of forage soiled by very 
active virulent matters, can also occasion the disease. 
In the case of cutaneous wounds, the characteristic 
tumor develops at the place of inoculation itself; in 
the two latter contingencies the hacilli multiply in the 
blood and determine tumors in places where they meet 
with an opening by which they may penetrate into 
the connective or muscular tissue. Rupture of some 
of the fibers of the muscles, alteration of the vascular 
endothelium (perhaps by the products of the bacilli), 
the production of even a slight wound which may be 
overlooked, are all so many factors on which depend 
the seat of the primary and secondary tumors. 

Attenuations. Preventive inoculations. The virus be- 
comes spontaneously attenuated when it is left in con- 
tact with the air; diminution of its virulence can be 
obtained artificially by means of antiseptics and heat. 
The latter agent supplied to MM. Arloing, Cornevin, 
and Thomas, the means of preparing virus of various 
degrees of intensity. The natural serosity of the 
specific lesions can be attenuated to different degrees 
by a temperature of from 65 to 70, maintained for a 
greater or less length of time. These authors, how- 
ever, operated by preference with serosity dried at the 
temperature of 30 to 35; the dried virus is, in reality, 
more fixed than the fluid serosity, because the spores 
are much more resistant when dry than when in a 
moist condition. This dried virulent substance with- 
stands temperatures of 80 to 90 without losing any 
of its activity; reduced to powder and moistened, 
then brought to temperatures varying between 60 and 
110, it becomes progressively attenuated. The virus, 
attenuated to such an extent that it is no more fatal, 



202 Manual of Veterinary Microbiology. 

vaccinates the organism against more active virus. 
The French authors recommend the successive em- 
ployment of two vaccines, one exposed to 100 
the other to 90, during seven hours. These two 
vaccines are dry when taken from the oven in 
which they have been prepared ; the dose employed 
is one centigram of the powder diluted in a gram of 
water for each animal. The vaccine prepared at 100 
is first used, and then, after eight days, the other. 
The inoculations are made in the cellular tissue of the 
ear, or in the internal face of the end of the tail. The 
autumn or the end of winter are the seasons selected 
for the operation. It is necessary to guard against in- 
oculation during very hot weather as an elevated 
temperature increases the activity of the virus. 

The consequences of the vaccinal inoculation are 
local and general. At the inoculated point an en- 
gorgement develops, generally of small extent and 
healing spontaneously. At the same time there is 
observed a febrile reaction, which indicates the exist- 
ence of the disease in a mild form. 

Kitt recommends a single vaccine prepared at 90. 

When the virus has been thus artificially attenuated 
it can regain its original activity by successive pas- 
sages through the bodies of young guinea pigs, and 
also under the influence of lactic acid, etc., as already 
described. 

Animals can also be vaccinated by injection of the 
natural virus either in the cellular tissue or into the 
blood. We have seen above, that inoculation of small 
doses in the connective tissue confers immunity in 
place of the disease, and that the blood tolerates com- 
paratively large doses of the virus. Injection into 



Microbic Diseases Individually Considered. 203 

the veins, however, must be made with the greatest 
care in order to avoid the accidental deposition of any 
of the virus in the surrounding tissue, hence, in prac- 
tice, the attenuated viruses are generally preferred to 
the natural viruses, for the production of immunity. 

Rouget of the pig.* 

This disease, peculiar to the pig, is infectious and 
contagious ; it chiefly attacks adult animals and those 
of the improved breeds. It manifests itself by a very 
intense febrile reaction, by red or purple patches, at 
first discreet and afterward confluent, upon the integu- 
ment, by a diarrhoea more or less intense, succeeding 
to constipation, and often by cough. The redness of 
the skin may be absent in very acute cases. 

The duration of the disease is always short, on an 
average, two days ; it may, however, last four or five 
days, and, on the other hand, may occasionally be al- 
most fulminating in character. 

Death is the usual termination, but a considerable 
proportion of pigs may recover. Moreover, all the 
the pigs which have been in contact with diseased 
animals do not necessarily contract the disease. 

The autopsy discloses a general congested condition 
of the capillaries. There is injection and serous infil- 
tration of the skin and subcutaneous cellular tissue, 
injection with petechiae and sero-fibrinous exudation 
of the peritoneum, pleura and pericardium, changes 
of the same kind in the gastro-intestinal canal in 
which the mucosa is reddened, thickened and infil- 

* [Erig. Swine erysipelas ; Ger. Rothlauf. The disease has not 
been recorded in the United States or Canada. D.] 



204 Manual of Veterinary Microbiology. 

trated ; in many places the epithelium is desquamated, 
and occasionally there are ulcerations in way of forma- 
tion. 

The blood vascular glands (spleen, lymph nodes, 
Peyer's patches) are tumefied by congestion, exuda- 
tion and extravasation ; the kidneys, lungs and heart 
always show injection and even extravasations. In 
rare cases there are multiple lesions of broncho-pneu- 
monia. 

The germ of rouget is a very fine, cylindrical bacil- 
lus recalling that of mouse septicaemia ; (1) it measures 
~\.p. to 2fjL in length by O-l// to O15// in thickness. It is 
found in'the blood, especially in the fine capillaries, in 
contact with their internal wall ; it also exists in the 
exudates and in all the diseased organs : liver, spleen, 
kidneys, lymphatic glands, marrow of bones, etc.; in 
the fsecal matters, and in the urine. This agent should 
not be confounded with a large rod which is found in 
the blood in most diseased pigs and which does not 

(1) Mouse septicaemia was obtained experimentally by Koch, by 
inoculating putrefied blood under the skin of these -animals. The 
disease is easily transmitted to house mice, whilst field mice are 
refractory. The microbes show much resemblance with those of 
rouget ; they have the same form, the same dimensions, are rather 
anaerobic than aerobic, and both take the Gram and Weigert 
stains. In gelatin, stab cultures of mouse septicaemia give colo- 
nies with radiating branches like those of rouget, but the rays are 
confounded with each other, whilst those of rouget are distinct ; 
gelatin is not fluidified by either. Potato is poorly or not at all 
adapted to their vegetation. Both become much elongated in 
cultures. The pigeon succumbs with equal rapidity to rouget and 
mouse septicaemia. The rabbit, on the other hand, is much more 
sensitive to the former of these diseases. Both produce, in the 
pigeon and the mouse, enlargement of the spleen and congestive 
lesions of the different organs. 



Microbic Diseases Individually Considered. 205 

show any specific pathogenic property. This last 
germ is never seen in the experimental disease and 
probably comes from a secondary invasion occurring 
in the diseased animal. The point of departure of 
this collateral infection appears to be the intestine, in 
which the bacillus just mentioned is found in abun- 
dance; it is present in the blood in smaller numbers 
the further removed the latter is from the abdominal 
cavity. 

Pasteur and Thuillier, who first described the bacil- 
lus of rouget, describe it as of a figure 8 form, but 
this was an error of observation which our staining 
methods and improved instruments have corrected. 

The bacillus of rouget is non-motile. 

Cultures. The bacillus of rouget is especially ana- 
erobic but it also grows in contact with the air ; it 
multiplies at a temperature as low as 20, but grows 
especially well in the incubator. 

The blood and the pulp of the various diseased or- 
gans may be used for the inoculation of culture media. 
This material will more likely be pure when taken 
from a part at some distance from the abdominal cav- 

ity- 

The germ grows well on the various culture media : 
on potato, especially in presence of oxygen, the growth 
is feeble. 

In bouillon it produces, after forty-eight hours, a 
slight uniform turbidity, which afterward becomes de- 
posited as a whitish gray sediment. 

Stab cultures in gelatin are characteristic. The 
germ multiplies especially in tho deeper parts and 
forms along the course of the puncture a track from 
which radiate silky tufts, giving to the whole the as- 



206 Manual of Veterinary Microbiology. 

pect of a bottle brush. This characteristic form is 
only obtained with gelatin of firm consistence. The 
bacillus of rouget appreciably lengthens in its cul- 
tures. 

Research and coloration. The specific germ can 
p. g be demonstrated in the blood, in the 

fluid exudates, and in sections of the 
1- various tissues affected; on account 
of its small size it is advisable to ex- 
Bacilli: 1, of amine only stained preparations and 
pneumo-enteritis to employ a magnification of 800 di- 
ofthepig;2,ofrou- ameters, at least. The bacillus of rou- 
get is stained by the hydro-alcoholic 
solutions of all the aniline colors ; the color is not re- 
moved by the reactions of Gram and Weigert. 

Experimental inoculations. The bacilli of rouget are 
fatal for the pig, rabbit, mouse and pigeon. The three 
last species are especially sensitive and succumb to 
subcutaneous, intra-peritoneal, or intra-venous inocu- 
lations of a virulent product, whether natural or de- 
rived from culture. 

Subcutaneous inoculation may fail in the pig ; in- 
gestion, although generally communicating the disease, 
is also uncertain in its results. The mouse and the 
pigeon succumb in two to four days; the pigeon takes 
the form of a ball of feathers, as in cholera ; the rab- 
bit dies in three to six days. The lesions noticed at 
the autopsy consist of a hypertrophy of the spleen 
with congestion of the liver and lymphatic glands. 

The guinea pig, rat, dog, and chicken are refractory 
to experimental inoculation. 

Etiology and pathogeny. Although some tests of in- 
fection by the digestive passage performed in Germany 



Microbic Diseases Individually Considered. 207 

by Lydtin were unsuccessful, there is reason to believe 
that, under natural conditions, the virus gains entrance 
by the digestive canal. 

The virulent matter ingested by healthy pigs will 
consist especially of the intestinal dejections of the 
diseased. Lack of proper attention to the feeding 
troughs, and defective conditions of the pens (ventila- 
tion, lighting, cleanliness) are of a nature to favor the 
propagation of rouget, as well as of other diseases of 
the pig. 

Adult hogs, especially those of the improved breeds 
(English), are most liable to contract the disease ; 
young pigs and those of native breed are more resist- 
ant. 

Attenuation. Vaccination. Rouget of the pig be- 
comes well acclimated in the pigeon and the rabbit, 
and in these two species acquires great virulence ; but 
whilst its repeated passage through the organism of 
the pigeon renders it more active for the pig, passage 
through the rabbit, on the contrary, diminishes its 
virulence for the pig. This attenuation, after a certain 
time, is such that the virus coming from the rabbit 
no more kills the pig; it, however, makes it sick and 
confers immunity on it against strong virus. The at- 
tenuation thus obtained persists in cultures afterward 
made in ordinary bouillon and these cultures can be 
used to vaccinate the pig. 

In practice two vaccines only are employed, and 
these in succession at ten days interval ; a feeble vac- 
cine is used first and then a second, the virulence of 
which is much stronger. 

Young pigs being much less susceptible to the dis- 
ease and therefore to the virus, the period of ,youth 



208 Manual of Veterinary Microbiology. 

should be preferred for vaccination. The immunity 
obtained lasts only one year, but this term is sufficient 
for the needs of breeding and fattening.* 

Pneumo-enteritis of the pig, hog cholera. 

Pneumo-enteritis of hogs is an infectious and con- 
tagious disease which was long confounded with 
rouget. It was described, in the first place, in Amer- 
ica, by Salmon under the name of hog cholera; in 
France it has been investigated by Riestsch and Jo- 
bert, and Cornil and Chantemesse, in connection with 
the epizootics at Marseille and at Gentilly; it was 
studied by Selander in the swine of Sweden and 
Denmark. 

This disease chiefly attacks young animals and is 
nearly always fatal. It manifests itself by symptoms 
the description of which differs somewhat in the dif- 
ferent countries in which it has been observed.^ 

According to Salmon, the disease may be acute or 
chronic. In the latter case inappetence is observed 
along with persistent diarrhoea and slow emaciation 
of the diseased animals. When the disease is acute 
the diarrhoea is more intense and sanguinolent. In 
both cases the intestine is much altered, principally 
the large intestine. The latter presents ulcerations 
and considerable thickening of its mucosa when the 
disease has been slow ; when the evolution has been 
rapid the lesions assume a hemorrhagic character and 
affect not' only the caecum and large colon, which are 

* [Lorenz has introduced a method of protective inoculation by 
the use of the blood-serum of swine which have previously been 
immunised against rouget. (Centralblatt fur Bacteriologie, xiii 11, 



Microbic Diseases Individually Considered. 209 

much injected and ulcerated, but also the spleen, 
liver, kidneys, and mesenteric glands. Generally the 
lungs are unaffected ; nevertheless, some foci of hepa- 
tization may be noticed in the last stage of the slow 
form of the disease. The skin of the neck and ab- 
domen, and sometimes of the whole body, is red- 
dened. 

Intestinal lesions, therefore, predominate in the dis- 
ease studied by Salmon. 

The disease studied in France is characterized by 
an intense fever with considerable prostration of the 
affected animals, staggering gait and sometimes paral- 
ysis. An intense and fetid serous diarrhoea soon 
supervenes ; this is often preceded, and occasionally 
followed, by constipation ; at the same time, or a little 
later, symptoms of pulmonary trouble become evi- 
dent : fitful hoarse cough, accelerated and embarrassed 
respiration, and abundant mucous discharge from the 
nostrils. 

The most conspicuous symptoms vary according as 
the intestinal or pulmonary troubles predominate. In 
the epizootic at Marseille enteritis was constant and 
the pulmonary lesions incidental ; on the contrary, 
broncho-pneumonia was the dominant feature in the 
epizootic at Gentilly. MM. Cornil and Chantemesse 
think that this peculiarity depends upon the mode 
of entrance of the virus. The Gentilly hogs were 
contaminated at the abattoir by inspiring air charged 
with virulent dust, whilst those of Marseille con- 
tracted the disease by ingestion of contaminated food. 

In the course of the disease a diffuse inflammation 
often develops on the skin of the lower part of the 
18 



210 Manual of Veterinary Microbiology. 

abdomen, on the perineum, groins, limbs, and at the 
root of the ears, these regions then taking a more or 
less pronounced red or violet color, and thus tending 
to increase the chances of confounding it with rouget. 
These cutaneous changes, however, are less constant 
than in the last disease. 

Pneumo-enteritis is of rather long duration : twenty 
to twenty-five days on an average, never less than 
eight to ten days ; it may extend to five or six weeks ; 
it is very contagious, and few hogs which have been 
exposed to the contagion escape. 

In very rapid cases the autopsy shows, beside ec- 
chymoses disseminated through the connective and 
inter-muscular tissue, peritoneum, pleura, pericardium, 
and heart, a violent inflammation of the stomach and 
intestines, with interstitial hemorrhages and erosions 
or ulcerations at Peyer's patches; the mesenteric 
glands are voluminous and infiltrated ; the lungs are 
normal or show lobular foci of hemorrhagic congestion. 
When the evolution of the disease has been slow the 
lesions are better defined; those of the intestine, 
csecum, and large colon are especially remarkable ; 
the wall of these organs is considerably thickened and 
indurated, and has become rigid. The swelling and 
induration chiefly affect the Peyer's patches ; these 
are the seat of a necrotic process which leads to the 
formation of grayish colored diphtheritic exudates and 
ulcerations of greater or less extent, both in area and 
in depth. The inflammation sometimes extends to 
the peritoneum. 

The lungs show lesions of broncho-pneumonia at a 
more or less advanced stage; pleurisy is also occasion- 
ally present. 



Microbic Diseases Individually Considered. 211 




The mesenteric and bronchial lymphatic glands are 
tumefied and sometimes partly caseous. 

Microbe. The 

germ of pneumo- Fi g- 9 - 

enteritis belongs to 
the group of bac- 
teria showing a clear 
central space, that 
is, those which take 
the stain better at 
their margins than 
in the center. It is 
ovoid, measures I/* 
to 2fj. in length by 
0-4// to 0-6// in thick- 
ness. It is motile, 
aerobic and facul- 
tatively anaerobic. 
It does not form spores. 

Action of physical and chemical agents. The bacillus 
of pneumo-enteritis is destroyed by a temperature of 
58, maintained during from fifteen or twenty minutes. 
It preserves its vitality in spite of desiccation for 
nearly two months. It vegetates and multiplies in 
water at the ordinary temperature of summer ; it re- 
tains its vitality for more than fifteen days in sterilized 
water. 

Those authors who have studied the action of chem- 
ical agents on this microbe especially recommend for 
its destruction mineral acids and sulfate of copper. 
MM. Cornil and Chantemesse recommend the follow- 
ing solution for the disinfection of pens and other in- 
fected objects : 



Hog cholera bacilli in spleen of guinea 
pig; cover-glass preparation. XI 200. D. 



212 Manual of Veterinary Microbiology. 

"Water . . . ^ . . 100 
Carbolic acid . * . . 4 
Hydrochloric acid ... 2 

Cultures Cultures on artificial media succeed well 
at temperatures varying from 18 to 45. 

Bouillon becomes turbid without showing any spe- 
cial characters. 

Gelatin, inoculated in superficial lines, shows a 
raised growth of white or bluish-white appearance, 
with irregular, often lace-like, borders ; the medium 
is not fluidified. Inoculated by puncture the culture 
shows itself in the form of rounded colonies covered 
with crystalline projections. 

The culture upon potato is remarkable for its clear 
brown color, which gradually becomes deeper with 
age. 

Sowings ought to be taken from the parenchyma- 
tous organs : liver, lungs, lymphatic glands, or from 
the blood. 

Schweinitz has isolated from cultures a toxic 
ptomaine (sucholo-toxin) and a special albumin (such- 
olo-albumin). 

Research and coloration. Salmon's bacillus readily 
takes up the different aniline colors ; simple methods 
of staining are alone applicable : hydro-alcoholic solu- 
tions, Loffler's blue, etc. The methods of Gram and 
Weigert, and Kiihne's violet completely fail. 

Experimental inoculations. The disease is inoculable 
to the mouse, rabbit and guinea pig. The pigeon 
takes it only from large doses. 

In the mouse the microbe assumes larger proportions 
than in the pig, and multiplies abundantly. 

The rabbit^ inoculated under the skin, succumbs in 



Microbic Diseases Individually Considered. 213 

from three to eight days according to the dose in- 
oculated ; the lungs are gorged with blood, the intes- 
tines are frequently the seat of a violent inflammation 
with resulting diarrhoea ; the spleen is tumefied and, 
along with the liver, often shows white necrosed foci. 
At the place of inoculation a creamy mass is found, 
also the product of a coagulative necrosis. 

The guinea pig dies in the same time, and with the 
same lesions, as the rabbit.* 

The pigeon is very resistant to pneumo-enteritis. It 
withstands small doses inoculated in the pectoral 
muscle although the inoculated point becomes the seat 
of a sequestrum similar to that produced by chicken 
cholera. Very large doses kill it in less than two 
days. 

The chicken is refractory. 

The pig is difficult to contaminate by hypodermic 
injection ; on the other hand, it succumbs ninety times 
out of a hundred to ingestion of virulent products. 
It contracts the same disease by inhalation of dust 
containing virulent matters held in suspension in the 
air, and also by intra-vascular injection. 

Etiology and pathogeny. The cause of pneumo-en- 
teritis resides in the bacillus discovered by Salmon ; 
the disease generally shows itself on a farm in conse- 
quence of the introduction of an infected hog ; it is 
transmitted from one hog to another by the ingestion 
of food or drink soiled by the intestinal dejections 
and nasal discharge of the diseased animals. 

* [The guinea pig appears to be somewhat less susceptible than 
the rabbit, and rarely shows the characteristic necrotic foci found 
in the liver in the latter animal. D.] 



214 Manual of Veterinary Microbiology. 

MM. Cornil and Chantemesse also conclude that the 
germ may penetrate by the respiratory passages and 
according to the way by which it is introduced, the 
disease will act more particularly upon the intestines 
or upon the lungs. 

According to Salmon, the natural virus is subject to 
considerable variations of activity, and this explains 
the sometimes rapid and sometimes slow course of the 
disease. 

The local lesions which follow its penetration assume 
the general character of an inflammation with a ten- 
dency to early mortification ; this tendency is suffi- 
ciently demonstrated by the diphtheritic exudates and 
ulcerations of the intestinal mucous membrane and 
the coagulative necrosis, under the form of caseous 
masses, in the glands of diseased hogs. This charac- 
ter also shows itself in the mouse, rabbit, guinea pig 
and pigeon which have been subjected to experimental 
inoculation. 

The local alterations lead to emaciation of the af- 
fected animals, but this erifeeblement is complicated 
with an intoxication. We have already seen that a 
toxic ptomaine has been isolated from cultures of 
pneumo-enteritis, and it may be that this poison, se- 
creted in greater abundance by very virulent germs, is 
the cause of the vascular changes met with in acute 
cases. 

The resistance of the bacteria to desiccation, and 
the facility with which they multiply in water at the 
ordinary temperature, are conditions which favor the 
persistence of the disease in one place and the produc- 
tion of new centers of infection. 

Attenuation. Vaccination. Attempts at attenuation 



MicroUc Diseases Individually Considered. 

of the bacillus of pneumo-enteritis have been made by 
MM. Cornil and Chantemesse. They had recourse to the 
action of heat upon cultures. A culture maintained 
at 43 during seventy-four days produces only a local 
abscess in the rabbit but is still regularly toxic for the 
guinea pig. After ninety days the virus no more 
kills guinea pigs ; they contract an abscess at the 
point of inoculation, whilst rabbits often escape even 
this lesion. The virus thus attenuated transmits its 
special virulence to its descendants and confers, on 
guinea pigs and rabbits which have received it, im- 
munity for virus which has been heated only seventy- 
four days; the latter acts in the same way toward 
more active and the natural virus. It is therefore 
possible to vaccinate the rabbit and the guinea pig 
against pneumo-enteritis. Unfortunately this method 
of prevention, applied to the pig, has not given the 
same result, and a process of vaccination is yet to be 
found for this species. 

The vaccine of rouget does not vaccinate hogs 
against cholera, a circumstance which increases the 
importance of the differential diagnosis. 

Schweinitz has succeeded in vaccinating the guinea 
pig by means of soluble substances which he has iso- 
lated from cultures.* 

* [Billings obtained protection against this disease by inocula- 
tion of pigs with cultures derived from mild cases of the natural 
disease. A certain proportion of the animals die from the inocu- 
lation. Neb. Ag. Exper. Sta., Vol. 2, No. 4. Selander (confirmed 
by Metschnikoff ) found that cultures of the Danish swine-pest, 
increased in virulence by passage through pigeons, produced in 
the blood of rabbits very active toxines. When such blood was 
sterilized at 57 C. and injected in rabbits it conferred a solid im- 
munity. The serum of rabbits thus vaccinated was also iound to 



216 Manual of Veterinary Microbiology. 

Pneumo-enteritis of the sheep. M. Galtier has studied 
a disease in the sheep to which, from its principal 
lesions, he has given the name of pneumo-enteritis, 
and which, according to this author, is caused by the 
germ of the disease of the pig which has just been 
de'scribed. This affection sometimes occurs in an epi- 
zootic form in sheep, and may make great ravages 
in affected flocks. In several cases the disease had 
originated in consequence of the introduction into the 
sheep-folds of swine recently purchased and which 
had contracted pneumo-enteritis in the market pens. 
.Once established in sheep, it transmits itself with great 
facility from sheep to sheep. 

The general symptoms consist in lassitude and gen- 
eral loss of vigor of the affected animals, with inappe- 
tence, loss of rumination and the appearance of a 
more or less intense fever. These symptoms are soon 
succeeded by bloating, fetid and exhausting diarrhoea, 
cough, accelerated respiration, mucous discharge from 
the nostrils sometimes streaked with blood, and the 
special symptoms of a broncho-pneumonia or of a 
broncho-pleuro-pneumonia. The skin and the visible 
mucous membranes take a more or less vivid red color, 
sometimes mixed with hemorrhagic points. In preg- 
nant females abortion is often observed although the 
mother does not necessarily succumb to the attacks of 
the disease. 

The disease may show various degrees of intensity ; 
it is sometimes very severe and kills in a few hours, or 
days, sometimes benign and passes unperceived. Con- 
possess immunising properties. Annales de Plnst. Pasteur, 1890, p. 
646. These results are not obtained with the American disease. 
(Smith & Moore.) D.J 






Microbic Diseases Individually Considered. 217 

valescence from the severe forms is always prolonged. 
The receptivity of sheep diminishes with age ; thus, 
the disease is noticed to be more severe and more fre- 
quently fatal in young animals. 

The bodies rapidly putrefy ; the subcutaneous and 
intermuscular connective tissue is dotted with hemor- 
rhagic points, sometimes with gelatinous exudates. 
The peritoneum, pleura, and sometimes the peri- 
cardium may be the seat of fibrinous inflammations. 
The mucous membrane of the fourth stomach, small 
and large intestine, is congested; it shows extrava- 
sated points and sometimes erosions ; Peyer's patches 
are tumefied. The liver is also hypersemic and dotted 
with petechise ; sometimes it contains abscesses when 
the disease has been somewhat prolonged. There are 
disseminated lesions of broncho-pneumonia with in- 
filtration and thickening of the interlobular connective 
tissue septa. The mucous membrane of the bronchi 
and of the trachea is reddened and thickened, and se- 
cretes an abnormal quantity of mucus. When the 
disease has developed slowly, caseous foci are not in- 
frequently found in the lungs. The lymphatic glands 
of the mesentery and of the root of the lung are en- 
larged, congested and infiltrated. 

The micro-organism which produces this disease of 
the sheep is identical, according to M. Galtier, with 
that of pneumo-enteritis of the pig. This author 
claims to have succeeded in transmitting this last dis- 
ease to the sheep by inoculation, as well as in trans- 
mitting the disease of the sheep to the rabbit, guinea 
pig,, dog, pig, goat, calf, to solipeds, to the chicken and 
to the pigeon. In the goat it resembles the pleuro- 
19 



Manual of Veterinary Microbiology* 

pneumonia (bou-frida) peculiar to this species ; in soli- 
peds it gives rise to symptoms recalling the typhoid 
disease; finally, it would explain epizootic abortion 
when it is spontaneously transmitted to cows. 

Transmission is readily procured by hypodermic, in- 
travenous and intra-pulmonary injection, less easily by 
the digestive canal. The virus loses its virulence by 
multiple transfers on artificial media and passages 
through individuals in which the disease develops 
slowly. On the contrary, its virulence increases in 
organisms very susceptible to its influence. Thus, ac- 
cording to M. Galtier, the pneumo-enteritis of the pigs 
at Gentilly is transmissible to sheep (contrary to the 
assertion of M. Nocard) when the substance inocu- 
lated is taken not from a culture but from a diseased 
animal. 

The natural contagion occurs by ingestion, and es- 
pecially by inhalation, of virulent products. The dis- 
ease is also transmitted from the mother to the foetus. 

According to the researches of M. Galtier, which 
we have just briefly reviewed, pneumo-enteritis, which 
is generally considered to be peculiar to the pig, ex- 
tends to all farm animals, especially to the sheep, 
bo vines and solipeds. The disease being transmitted 
to the foetus, calves coming from diseased cows which 
are or have been subject to coughing are born with 
the germ of the disease in them and die in a few days 
with the lesions of broncho-pneumonia and enteritis 
(pneumo-enteritis of calves). 

Infectious pneumonia of the pig, swine plague. 
This disease of swine has been described in Ger- 
many under the name Schweineseuche, and in Amer- 



Microbic Diseases Individually Considered. 

ica under that of Swine-plague. It is an infectious 
and epizootic disease, characterized by the predomi- 
nance of the pulmonary lesions. These consist of 
broncho-pneumonic foci with or without complications 
of pleurisy, enteritis, etc. The lung often contains 
caseous masses ; sometimes it is gangrenous. The 
evolution is acute or chronic ; in the first case the 
duration of the disease is from three to nine hours on 
an average. 

There is present, in the pulmonary lesions, pleura, 
peritoneum and pericardium, a non-motile microbe, 
ovoid, from 1/z to 1-2/* long by 0.6/z in thickness, stain- 
ing only at its extremities. 

The disease is inoculable to the mouse, rabbit, 
guinea pig, chicken and pigeon, but large doses are 
required to produce fatal results in the last three 
species. 

It is transmitted from one hog to another by inhala- 
tion, probably also by ingestion, and perhaps by acci- 
dental cutaneous inoculation. 



220 



Manual of Veterinary Microbiology. 



Differential diagnosis of rouget, pneumo-enteritis, and 
swine-plague (jieste-porcine.)* 



BOUGET. 


PNEUMO-ENTERITIS. 


SWINE PLAGUE. 


Course rapid: 2 to 5 


Course slow ; 20 to 25, 





days. 


days, never less than 8 to 






10 days. 




Attacks especially adult 


Attacks especially young 




hogs. 


pigs. 




General symptoms pre- 
dominating". 


Gastro-pulmonary symp- 
toms predominating. 


Pulmonary localiz 
tion predominating. 


Redness more extend- 


Redness less extended, 




ed, more constant. 


less constant. 




Congestive lesions o f 
all the organs, with pe- 


Inflammatory lesions of 
a necrotic character in the 


Broncho-pneumonia 
with caseous foci. 


techial extravasations 


intestine posterior to the 




and exudative inflamma- 


ileo-co3cal valve, and in the 




tions of serous mem- 


mesenteric and bronchial 




branes. 


glands. 




In very slow cases the 


When the disease is rapid 




intestine may contain ul- 


the intestinal changes may 




cerations at Peyer's 


not have reached their 




patches as in pneumo-en- 
teritis. 


usual stage and thus may 
cause the disease to be mis- 






taken for rouget. This mis- 




' i * ~ 


take is more easily made 







in cases in which the pul- 






monary lesions have not 






had time to develop. 




Bacilli cylindrical, non- 


Bacteria ovoid, motile, 


Bacteria ovoid, non- 


motile, stained by the 


not stained by the Gram or 


motile, staining only 


Gram and Weigert meth- 
ods. 


Weigert methods. The cen- 
ter stains much less than 


at their extremities. 




the periphery. 




Especially anaerobic. 


Especially aerobic. 




Inoculable to the mouse, 


Inoculable to the mouse, 


Inoculable to the 


rabbit and pigeon, but 


rabbit, and guinea pig, to 


mouse and to the rab- 


not to the guinea pig. 


the pigeon only when very 


bit, and when large 




large doses are employed; 
not to the chicken. 


doses are employed to 
the guinea pig, pigeon 






and chicken. 


Culture in gelatin has 


Culture has the form of 




the appearance of a test- 


globules covered with crys- 




tube brush. 


talline asperities. 





* [From the lack of distinctive names for the various infectious 
diseases of swine which have been described in different coun- 
tries, the morphological and biological similarity of their germs, 
and the absence of any post-mortem lesions which are absolutely 
characteristic, there still exists considerable confusion as to their 



Microbic Diseases Individually Considered. 221 

Tuberculosis. 

Tuberculosis is a disease too well known to require 
a description of its symptoms and lesions in this 
place. We will conside'r it only from the special point 
of view of its bacteriology. 

The disease produces its greatest ravages in the hu- 
man species, and then, with a frequency decreasing in 
the order in which they are named, it attacks the 
bovine species, the pig, horse, dog, and cat. Fowls 
also are decimated by this terrible scourge. The 
question as to the identity of human and avian tuber- 
culosis has been much discussed and is still in dis- 
pute. 

The distribution of the specific alterations is some- 
what different in the different species. 

In cattle they are most frequently found in the 
lungs, pleura, and thoracic glands, but are also com- 
mon in the intestines, peritoneum, liver, kidneys, 
mammae, and the corresponding lymphatic glands; 
they have also been met with in the meninges, inter- 
identity. The names given in each of the lists below seem to re- 
fer to the same disease : 

I. Rouget, in France ; rothlauf , in Germany ; swine erysipelas, 
in England (cases described by McFadyean, Jour. Comp. Path., 
Vol. IV, Part 4). 

II. Pneumo-enteritis (Klein), swine fever, in England ; svinpest 
(Selander), in Denmark ; hog cholera (Salmon and Smith), swine 
plague (Billings), in America. 

III. Schweineseuche, in Germany ; swine plague (Smith), in 
America (uncertain). 

The germ of the French disease of hogs described as pneumo- 
enteritis, as well as that of Galtier, has not been identified with 
that of either of the diseases included under lists II and III. D.] 



222 Manual of Veterinary Microbiology. 

muscular connective tissue, bones, bone marrow, and 
the articulations. The disease is especially frequent 
in adult animals; when it exists in the calf it is lo- 
cated nearly always in the abdominal viscera, and 
more especially in the liver when the affection is con- 
genital ; but it can quickly extend to the thoracic or- 
gans. We have had the opportunity of establishing 
this extension to an excessive degree in the viscera 
coming from the abattoir of Brussels, the lungs and 
bronchial glands showing a generalization of miliary 
tubercles. In two cases of intra-uterine infection ob- 
served by MM. Malvoz and Brouwier the lung was 
exempt from lesions, these occupying the liver, hepatic, 
and bronchial glands. 

The pig, although very susceptible to the experi- 
mental disease, appears to be rarely affected with tu- 
berculosis ; here it is the pulmonary form that pre- 
dominates. M. Moule has communicated a case in 
which he observed extension to the pleura, ribs, and 
muscles. Microscopical and bacteriological researches 
have enabled us to connect with tuberculosis those 
scrofulous alterations of the cervical glands which are 
occasionally observed in the pig. According to M. 
Nocard, the disease in this species often develops with 
great rapidity and passes unperceived ; in the chronic 
forms the bacilli are rare and seem to have lost part 
of their virulence; inoculated to guinea pigs they pro- 
duce a disease of slow course ; but the period of in- 
cubation becomes shortened again when these bacilli 
are inoculated from the first guinea pig to others in 
series. This property belongs also to the lesions of 
human scrofula. 

In the horse two forms of tuberculosis exist ; the 



Microbic Diseases Individually Considered. 223 

abdominal form, the most frequent, is characterized 
by confluent lesions in the spleen, mesenteric glands, 
liver, and intestines; in the thoracic form the changes 
occur chiefly in the lungs ; the latter also develop, but 
more slowly, as a sequel to abdominal tuberculosis. 
According to M. Nocard, who gave us our first in- 
formation upon tuberculosis of the horse, this disease 
is often accompanied with a polyuria of remarkable 
intensity. 

In the dog the disease also occurs under the two 
forms observed in the horse. A considerable number 
of cases of tuberculosis in this species have already 
been communicated. We ourselves have seen two 
cases. In the first the liver was enlarged and infil- 
trated with a neoplastic tissue of a grayish color, which, 
under the microscope, was resolved into miliary tuber- 
cles destitute of giant cells. The hepatic and mesen- 
teric glands were much tumefied, and partly caseous. 
The lung contained two nodules of the size of a pea. 
The second dog showed generalized tuberculosis of 
both lungs, and of the bronchial glands. 

The ape is very susceptible to the disease, and very 
readily contracts it in our climate. 

In fowls the abdominal viscera are most affected, 
and this often to an excessive degree. In this species 
the liver seems to be the -place of predilection for the 
tubercles ; sometimes the latter are absent, only a con- 
siderable enlargement with degeneration of this or- 
gan being observed, but the spleen, intestines, and 
peritoneum may, at the same time, be studded with 
lesions. The lung is rarely affected. 

Microbe. The efficient cause of tuberculosis resides 
in the bacillus of Koch. This shows itself under the 



224 Manual of Veterinary Microbiology. 

form of a homogeneous rod, or more frequently com- 
posed of granules arranged in linear series. It is 
straight or "bent in the form of an arc, sometimes 
S-shaped; it measures 2fj. to 6// in length by 0-3/z to 
0-5, in thickness. The rod is uniform in size through- 
out its extent. 

According to Cornil and BabSs the granular appear- 
ance is especially observed in bacilli long abandoned 
to the air ; these authors have demonstrated the same 
granules in bacilli coming from cultures and they re- 
gard them as spores. 

A certain number of the bacilli are 
^ observed to become considerably elon- 

"/ \f^~L gated and to swell at one of their ex- 
tremities; we have often seen these 
Tubercle bacilli, abnormal forms in bouillon cultures 

(M. and L.) of ayian tuberculosis. 

Action of physical and chemical agents. Tubercle 
bacilli are killed by a temperature of 70, maintained 
during ten minutes (Yersin). According to M. Gal- 
tier, heating at 71 during ten minutes does not suffice 
to sterilize tuberculous matter. Moist heat at 100 
sterilizes it with certainty in a few minutes, but this 
is not the case when the virulent substance is in the 
dry state, for the dried spores withstand 100. 

The tubercle bacillus withstands freezing, putrefac- 
tion and desiccation. This last operation, when it 
occurs at temperatures near to 30, is, indeed, an im- 
portant means of preserving it. The bacillus retains 
its vitality for a considerable time in sterilized water 
(seventy days). 

According to Yersin, carbolic acid, at 5 per cent, 
kills the bacillus in half a minute ; sublimate, at 1 to 



Microbic Diseases Individually Considered. 225 

1.000, in ten minutes ; absolute alcohol in five minutes; 
but contact with these agents must be much more pro- 
longed in the case of tuberculous substances, sputa, 
etc. Hence, for the destruction of the virulence of 
these substances, moist heat should be preferred. 

The tubercle bacillus resists the action of the gastric 
juice. . 

Cultures. Vegetation of the bacillus on artificial 
media is not easily obtained ; the operation, however, 
is only delicate for the first generation. The culture 
media most favorable for this bacillus are those which 
contain an addition of peptone, glycerin, and even of 
glucose, in proper proportions. The incubating oven 
should be kept at a temperature in the neighborhood 
of 39, that temperature being best suited for the 
growth of the bacillus; multiplication is impeded by 
slight deviation from this temperature, and ceases 
altogether at about 35. As the germ is aerobic, the 
culture medium should also be freely exposed to the 
air. The sowing should be taken from virulent 
products from young tuberculous animals in which 
the evolution is rapid, and should be implanted at 
once upon serum ; it is transferred several times upon 
this medium before trusting it to others. 

On serum, at the end of twelve to fifteen days, 
small, round, whitish grains appear, and slowly in- 
crease in size ; these grains are slightly raised, dry, 
and scaly in appearance ; their growth is very limited 
in the first cultures and they become confluent only 
after the fourth or fifth generation ; vegetation is then 
more rapid and the whole surface of the serum be- 
comes covered with a thin, dry film, studded with ver- 
rucose prominences. 



226 Manual of Veterinary Microbiology. 

Sowing of tuberculosis of mammals directly upon 
agar failed of results in the hands of MM. Straus and 
Gamaleia. and even transference of serum cultures 
on to agar only succeeded well after four or five pas- 
sages on the serum. 

The bacillus, therefore, requires to become accli- 
mated on an artificial medium in order to obtain a 
vigorous growth. If it is then transferred to animals 
it vegetates satisfactorily. 

The appearance of cultures on agar resembles that 
of serum cultures. 

In the case of avian tuberculosis much richer cul- 
tures are obtained directly upon serum. These cul- 
tures begin by rounded, whitish spots, waxy and 
moist, which after a few transfers produce a continu- 
ous layer of the same appearance, thus contrasting 
with the meager and dry film of human tuberculosis 
(Straus and Gamaleia). Cultures on agar and on 
bouillon are also more readily obtained and more 
abundant than in the case of human tuberculosis. 

In bouillons, after a few days, small flakes appear 
which gradually increase in size and fall to the bot- 
tom of the liquid without becoming dissociated ; they 
break up into finer particles only when the vessel is 
shaken. 

The tubercle bacillus also vegetates on potato, al- 
though this is not a very favorable medium for its 
growth. 

Research and coloration. Koch's bacillus fixes color- 
ing matters with difficulty and hence requires pro- 
longed exposure. In order to abbreviate the opera- 
tion recourse is often had to heat. The coloration, 
however, if slowly obtained, is persistent even against 



Microbic Diseases Individually Considered. 227 

the action of strong acids nitric and sulphuric ; upon 
this property are based the different methods of double 
staining which we will now describe. This property 
is also possessed by the bacillus of leprosy. 

1. Ehrlicfts method. 

The staining fluid is composed as follows : 

Aniline water (1) . . -9 cub. cent. 
Absolute alcohol .... 1 cub. cent. 
Concentrated alcoholic solution of 
fuchsin, methyl violet, or gentian 
violet .1 cub. cent. 

Cover glasses remain in this solution half an hour 
at least, sections twenty-four hours. 

Decoloration is obtained by a dilution of nitric 
acid 1 part nitric acid in 5 parts distilled water or in 
10 parts alcohol. Cover glasses or sections should re- 
main in the decolorizing fluid only from half a min- 
ute to a minute. They are then washed in distilled 
water and mounted. If it is desired to stain the 
background of the specimen so as to render the color 
of the bacilli more distinct, the preparations, after 
coming from the distilled water, are placed in a hydro- 
alcoholic solution of methylene blue if fuchsin has 
been the first stain, or in eosin, safranin, bismark brown, 
etc., if the bacilli have been stained violet ; then they 
are mounted. 

The time required by this method may be abridged 
by raising the temperature of the staining fluid to 

(1) Aniline water is made in the following manner: A drop of 
aniline oil is added to a few cubic centimeters of distilled water in a 
test tube ; the tube is vigorously shaken in order to dissolve the 
oil, and a few drops of absolute alcohol added to complete this 
solution. 



"228 Manual of Veterinary Microbiology. 

near the boiling point before the introduction of the 
specimens to be stained. The necessary staining is 
thus obtained in a few minutes. 

2. Lubimoff's method. 

The staining fluid is composed of: 

Water 20 cub. cent. 

Boric acid . . . 0-5 grams. 

Absolute alcohol . . 15 cub. cent. 
Fuchsin .... 0*5 grams. 

This fluid keeps indefinitely. 

To stain the bacilli upon the cover glass a few drops 
of the solution are deposited upon its surface and 
heated for two minutes over a spirit lamp; the 
preparation is then rapidly decolorized in sulfuric 
acid diluted to 1 to 5, then rinsed with alcohol and 
immersed in a concentrated alcoholic solution of 
methylene blue ; it is then washed with water, dried 
and mounted in balsam. 

Sections of tubercular tissue are left during one or 
two minutes in the staining fluid, previously heated 
to near the boiling point, then passed for a few seconds 
into alcohol and thence, for one to two minutes, into 
a 1 to 5 dilution of sulfuric acid, again into alcohol 
and, finally, for one minute, into a hydro-alcoholic so- 
lution of methylene blue. Dehydration is accom- 
plished by passing the sections through absolute alco- 
hol and xylol and they are then mounted in balsam. 

3. Ziehl-Neelsen method. 

The staining fluid is composed by mixing : 
Fuchsin . 1 gram. 

Absolute alcohol 10 grams. 

Five per cent aqueous solution of car- 
bolic acid. . . .-.-: . 100 grams. 



Microbic Diseases Individually Considered. 229 

The different steps of the operation are exactly the 
same as in LubimofPs method. 

4. Herman's method. 

The staining fluid is made extemporaneously and 
from two solutions : 

(1) A one-per-cent aqueous solution of carbonate 
of ammonia; 

(2) An alcoholic solution of methyl violet 6 B (1 of 
the violet to 30 of 95 per cent alcohol). 

A few drops of the second solution are added to 
several cubic centimeters of the first until the mixture 
obtains a deep violet color and this is brought to a tem- 
perature approaching ebullition. The cover glasses or 
sections are left in the stain from one to two minutes, 
then placed for from two to three seconds in nitric 
acid diluted 1 to 4 for sections and 1 to 10 for cover 
glasses ; after passing through the dehydrating fluids 
they are ready for mounting. 

Double staining may be obtained by immersing the 
preparations, after passing through the nitric acid and 
the alcohol, in an aqueous or alcoholic solution of eosin. 

5. Kitt's method. 

This requires two fluids both of which preserve 
themselves indefinitely : * 

(1) Aniline water, . . . . 100 grams. 
1 per cent solution of caustic soda, 1 " 
Fuchsin, . . . . 4 to 5 " 

(2) Alcohol, 50 grams. 

Water, . .... 30 u 

Nitric acid, . T . . 20 " 

Methylene blue to saturation. 

* [The aniline-containing solutions will generally be found unfit 
for use after one or two weeks. D.] 



230 Manual of Veterinary Microbiology. 

The preparations, cover glasses or sections, are left 
for from two to five minutes in the fuchsin solution 
heated as in the preceding methods, and then trans- 
ferred to the second solution, in which they remain 
one to two minutes. During this last period all the 
elements other than the Koch bacilli are decolorized 
under the influence of the nitric acid, but, at the same 
time, fix the blue so that double staining is combined 
with decoloration. 

The preparations are washed, dehydrated, and 
mounted. 

All these methods have the same value in so far as 
the coloration which they give is concerned. In prac- 
tice, however, those solutions may be recommended 
which are of longest preservation, and which require 
the fewest manipulations. The method of Kitt es- 
pecially fulfills these two conditions.* 

In the living subject about the only materials available 
for examination are fluid products, such as nasal dis- 
charge, milk or pus. In examining milk for tubercle 
bacilli the cover glass, after drying and before staining, 
should be freed from fatty matters by immersion in a 
mixture of absolute alcohol and ether, or in chloro- 
form. If a deposit is formed at the bottom of the 
liquid this will furnish the best material for examina- 
tion. 

Very often the Koch bacillus has to be sought for 
in cadaveric products. In this case the investigations 
will be directed to the tubercular lesions or those sus- 

*[0f the two methods most frequently employed Ehrlich's 
and Ziehl's the latter, on account of the greater preservation 
of the staining fluid, is often the most convenient, whilst the 
former gives a more brilliant color to the bacilli. D.] 



Microbio Diseases Individually Considered. 231 

pected from their physical characters to be of a tuber- 
cular nature. 

In the case of young, gray tubercles it suffices to 
spread upon a cover glass the product obtained by 
scraping their cut surface: the bacilli are here uni- 
formly distributed. But when the tubercles are al- 
ready caseated the caseous matter frequently contains 
but few bacilli except in birds, where, on the contrary, 
they are very numerous; this substance is first re- 
moved and the bacilli sought for in the wall of the 
caseous tubercle. The same method may be employed 
for cavities although these often contain a liquid very 
rich in bacilli. 

Experimental inoculations. Tuberculosis is inocula- 
ble to the horse, ass, ox, sheep, pig, dog, cat, rabbit, 
guinea pig, and to fowls. 

Subcutaneous inoculation is without effect in the 
horse, ass, sheep, pig, dog, cat, and chicken. 

Ingestion of virulent substances produces the dis- 
ease, but not in all cases, in the horse, sheep, pig, dog, 
and cat. Cattle are easily tuberculized in this way. 
The chicken remains unaffected when sputa or tuber- 
cular products of mammals are mixed with its food. 

Intravenous injection gives much surer results. 
Except in the case of birds it is almost invariably fol- 
lowed by a generalized tuberculosis of the lung with 
possible extension to* other organs. In the ass the 
experiments of M. Chauveau have shown that these pul- 
monary granula heal spontaneously after a few weeks. 
Fowls do not usually contract the tuberculosis of 
mammals by way of the circulation. Thus, MM. 
Cadiot, Gilbert, and Roger have recently announced 
that out of forty pullets inoculated by them either in 



232 Manual of Veterinary Microbiology. 

the veins or in the peritoneum, five only developed 
tubercular lesions. 

The goat forms a specially unfavorable field for the 
development of tuberculosis and was long considered 
to be absolutely refractory to the inoculated as well as 
to the spontaneous disease. M. Nocard has recently 
described the evolution of the disease in a goat inocu- 
lated in the jugular five years before, and which at 
last had become affected with mange ; M. Colin has 
also produced the disease in a goat inoculated under 
the skin with particles from the ox. 

The laboratory animals guinea pigs and rabbits 
are very susceptible to the disease. The guinea pig 
is endowed with a quite special receptivity which 
makes it the reagent par excellence for tuberculosis. 

Subcutaneous inoculation on the internal face of 
the thigh in the guinea pig is followed by a local ab- 
scess when the inoculated substance contains at the 
same-time pyogenic germs, or only by a few yellowish 
granulations if it is pure ; at the end of ten to fifteen 
days there -supervenes an engorgement, sometimes an 
abscess, of the superficial inguinal lymphatic glands; 
the sublumbar glands of the corresponding side are 
invaded about the twentieth day ; between the twenty- 
second and twenty-fifth days tubercles appear in the 
spleen and retro-hepatic glands ; the lungs, liver, and 
the other lymphatic glands are attacked later. The 
disease lasts about two months. When the inocula- 
tion has been made at the ear the invasion takes place 
by the anterior lymphatics, and the lungs are attacked 
before the abdominal viscera. 

Subcutaneous inoculation on the internal face of 
the thigh or at the ear, in the rabbit, does not give 



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Microbic Diseases Individually Considered. 233 

rise to engorgement of the corresponding lym- 
phatic glands. The local lesion is less pronounced 
than in the guinea pig, and generalization, less con- 
stant, occurs through the intermediation of the blood ; 
the changes, here, are found chiefly in the lung. 
With bovine tuberculosis, in certain cases in the rab- 
bit, M. Arloing has seen the development of glandular 
lesions resembling those of the guinea pig. 

Intra-peritoneal inoculation in the rabbit and guinea 
pig determines tubercular lesions of the peritoneum, 
epiploic glands, liver, and spleen. The duration of 
the disease is always shorter than by the subcutaneous 
method. 

Intra-vascular inoculation produces a generalized 
tuberculosis, but death is so rapid (fifteen to twenty 
days) that the specific lesions are not visible to the 
naked eye. The bacilli are disseminated throughout all 
the parenchymatous organs (septicsemic type, Yersin). 

According to Straus and Gamaleia this tuberculous 
septicaemia is obtained only with the cultures of avian 
tuberculosis. 

Diagnosis of doubtful cases in cattle. In cattle the 
diagnosis of tuberculosis by means of the clinical 
symptoms is often difficult. Not to speak of abdom- 
inal forms which are still more difficult of recognition, 
the discharge from the nostrils is often absent in pul- 
monary tuberculosis, and the search for the essential 
element the bacillus consequently denied to us. 

M. Nocard, basing himself on the fact that cattle 
are accustomed to swallow their expectorations, ad- 
vises looking for the bacillus in the pharyngeal mu- 
cus ; this can be obtained by scraping the mucous 
20 



234 Manual of Veterinary Microbiology. 

membrane of the throat with a spatula. Cagny pro- 
poses, in order to increase the bronchial secretion, the 
injection under the skin of ten to twenty centigrams 
of veratrine. 

Poels has resorted, in the absence of discharge, to 
tracheotomy and the examination of the tracheal 
mucus. 

The rarity of tubercular lesions of Demour and 
Decemet's membrane tends to refute the assertion 
of M. Mandereau as to the constant presence of the 
bacillus in the aqueous humor of tuberculous animals, 
and this assertion, which promised an easy diagnosis 
of the disease, has quickly been contradicted by Le- 
clainche and Greffier, whose investigations on twenty 
animals which were certainly affected always gave 
negative results. 

M. Peuch, having placed an -irritating seton in a 
tuberculous cow, found, by inoculating the pus of the 
seton to guinea pigs, that the bacilli passed into the 
pus from the eighth to the fourteenth day. He there- 
fore recommends the application of such an exudatory, 
and subsequent inoculation in order to remove the 
uncertainty of the diagnosis. 

If the presence of the bacillus in the expectoration 
enables us to affirm the existence of the disease, its 
absence does not authorize us in positively affirming 
that it does not exist. Sometimes it is necessary to 
resort to inoculation of expectorated products, and 
the same precaution should be observed when dealing 
with milk, etc. Under such circumstances we have 
recourse preferably to the guinea pig, and only in de- 
fault of this animal, to the rabbit. 

When we possess pure products cultures or young 



Microbic Diseases Individually Considered. 235 

tubercles collected in a state of purity and reduced to 
pulp the inoculation can with advantage be made 
into the peritoneal cavity, thus obtaining a more 
rapid evolution ; when we have only at our disposal 
virulent matters contaminated with other germs, such 
as pus or nasal discharge, we must content ourselves 
with subcutaneous inoculation. A region should be 
preferred in which the lymph glands are easily ex- 
plorable in order that the progress of the lesions may 
be followed. In default of this exploration the ema- 
ciation of the subject after the time specified above 
will enable us to affirm, before the autopsy, that the 
inoculation has been successful. 

Tuberculin. Another means of diagnosis consists in 
testing with Koch's lymph or tuberculin. 

Cultures of the tubercle bacillus contain a soluble 
product discovered by Koch which possesses a very 
remarkable property. This substance is without effect 
on healthy individuals, whilst it is toxic for the tuber- 
culous. The name tuberculin has been given to a 
glycerin extract of cultures which contain this active 
agent. Since his first discoveries Koch has simplified 
the process by which it is prepared. Large cultures 
in veal bouillon, to which has been added one per cent 
of peptone and from four to five per cent of glycerin, 
are reduced to one-tenth their volume at a tempera- 
ture of about 100, then filtered through porcelain so 
as to remove all the microbes. The tuberculin thus 
obtained contains enough glycerin for its own preser- 
vation. The product has not a stable composition. 
Koch recommends testing its activity on tuberculous 
guinea pigs. According to this author, a good speci- 
men of tuberculin, in the dose of one centigram, kills 



236 Manual of Veterinary Microbiology. 

a guinea pig which has been inoculated eight to ten 
weeks before ; it requires twenty to thirty centigrams, 
sometimes even as much as fifty centigrams, to kill a 
guinea pig inoculated four to five weeks before. The 
guinea pigs die in six to thirty hours, according to 
the extent of the tubercular process. 

Tuberculin, after twenty-four hours' treatment with 
two to three volumes of alcohol, yields up its active 
substance in the form of a precipitate of an albuminoid 
nature. 

Tuberculin causes in tuberculous subjects : 

1st. A more or less intense febrile reaction which 
supervenes after several hours, usually from the tenth 
to the twentieth ; 

2d. A quite remarkable inflammatory reaction 
around the tuberculous foci. This substance is there- 
fore pyretogenic and phlogogenic, but as the first of 
these actions produces its effects indirectly it is some- 
what delayed. According to Gamaleia, the mode of 
action of tuberculin is somewhat as follows : It pos- 
sesses properties which are especially toxic for the ele- 
ments of the tubercle, cause them to undergo necro- 
biosis in the same way as the secretions of the bacillus 
contained in the lesions produce necrobiosis (under 
the form of caseation or softening) of the central parts 
of the latter. Now, the proteins resulting from the 
decomposition of the elements thus attacked excite a 
local exudative inflammation and leucocytic infiltra- 
tion. This local reaction results in the breaking up 
and elimination of the tubercular foci. The febrile 
reaction must be attributed to the absorption of the 
necrosed tissues. (See page 64, Gangolphe and 
Courmont.) 



Microbic Diseases Individually Considered. 237 

The special hyperthermic action of tuberculin on 
tuberculous animals renders it a diagnostic agent for 
tuberculosis. 

The doses employed by different .experimenters 
have varied within very large limits; in general, the 
injection of 20 to 40 centigrams of tuberculin is suf- 
ficient. .From the tenth to the twentieth hour, oc- 
casiojially sooner, an elevation of temperature "of from 
one to three degrees is observed in tuberculous ani- 
mals. This is therefore an excellent means of bring- 
ing to light qbscure cases of tuberculosis. Unfor- 
tunately the rule is not without exceptions: some 
tuberculous subjects do not react at all and a certain 
number of others, not tuberculous, give the charac- 
teristic reaction. In spite of these exceptions tuber- 
culin still furnishes us with a supplementary means 
of diagnosis and ought not to be discarded. M. 
Nocard has shown that it has no injurious action on 
lactation or gestation ; he recommends its employ- 
ment in the sanitary inspection of dairies where 
milk is produced which is intended for public con- 
sumption.* 

On account of its phlogogenic and destructive ac- 
tion it might be supposed that tuberculin would act 
as a curative agent, but, unfortunately, this hypothe- 

* [Tuberculin during the last three years has been very exten- 
sively tested in Europe and in America and the results obtained 
are still entirely in harmony with the opinion here expressed. 
Animals in the last stage of the disease do not react; those which 
from any cause have an abnormal temperature at the time of the 
inoculation are also unsuitable. When the test is carefully made 
the result is almost always reliable, although the extent of the 
thermal reaction gives no indication of the extent of the diseased 
process in the animal. (D.)] 



238 Manual of Veterinary Microbiology. 

sis has not been confirmed; the numerous tests 
which have already been made have shown that tuber- 
culin, instead of exerting a curative effect, may be 
positively harmful and cause the extension if not the 
generalization of the disease. The reagent is, in 
reality, without effect on the bacillus, whilst the in- 
flammatory reaction which develops around the 
tubercles causes an aggregation of leucocytes which, 
becoming charged with microbes, transport these to 
points outside of the original lesion where they can 
incite new centers of disease ; this inflammatory re- 
action can, further, become directly harmful when 
the tubercles are numerous and occupy a large ex- 
tent of an important organ. 

Etiology and pathogeny. Contamination with tuber- 
culosis is most frequently indirect, but may also take 
place in a direct manner. 

Instances in which physicians and veterinarians 
have contracted the disease in making autopsies of 
diseased men or animals are incontestible, though 
fortunately rare. The virus inoculated through a 
wound, in such cases, occasions first of all a more or 
less limited cutaneous tuberculosis which may later 
become generalized. 

Transmission of the disease from mother to fcetus 
constitutes another example of immediate contagion. 
This mode of transmission, although now placed be- 
yond doubt both for animals and mankind, is actu- 
ally of rare occurrence; the tubercle bacillus is in 
reality confined to the specific lesions and only ex- 
ceptionally circulates in the blood ; moreover, it has 
not yet been demonstrated that it is capable of pass- 
ing through the villosities of the intact chorion, 



Microbic Diseases Individually Considered. 

intra uterine propagation appearing rather to depend 
upon some tubercular alteration of the maternal 
placenta. Johne has noted the existence of bacillar 
lesions in the liver and lung of a foetus found in a 
phthisical cow. MM. Malvoz and Brouwier have 
communicated two cases of congenital tuberculosis 
in the calf; the first of these cases is quite convinc- 
ing: the foetus was removed from the healthy womb 
of a cow affected with the generalized disease; the 
second describes the case of a calf six weeks old, the 
origin of which was not determined, but in which 
the lesions were regarded by the authors as congenital 
because they were located in the same organs as in 
the first case, that is, in the liver, and hepatic and 
bronchial lymph nodes. From the absence of intes- 
tinal and pulmonary lesions it was inferred that the 
infection could only have taken place through the 
umbilical vein. 

The presence of the bacillus having been demon- 
strated in the semen, some authors have been led to 
believe in the direct transmission from father to off- 
spring by infection of the ovum. The special locali- 
zation in the liver in well observed cases of congenital 
tuberculosis contradicts this manner of view. 

The tubercular virus may be directly transmitted 
from a diseased to a healthy individual through sex- 
ual intercourse, either from the female to the male, 
or inversely. 

Tuberculosis is usually communicated indirectly. 
The virulent matters rejected by the diseased (sputa 
of phthisical patients, nasal discharge and excrements 
of animals) and deposited on the ground, becoming 
dried and pulverulent, are carried with the air into 



240 Manual of Veterinary Microbiology. 

the respiratory passages of healthy individuals, or 
are deposited on their food. Sputa or nasal discharge 
may also be directly ingested by animals ; many in- 
stances of the propagation of human tuberculosis to the 
dog in this way have been recorded and it is probable 
that in the stables a considerable number of cattle 
contract the disease by consuming forage directly 
soiled by the expectorations of their neighbors. 

The milk of phthisical cows appears also to be an 
important carrier of the germ ; there is a difference 
of opinion on the question whether or not the udder 
can allow the passage of bacilli into the milk without 
itself becoming invaded by the tubercular process; 
some writers, basing themselves upon the result of 
numerous tests, have come to the conclusion that this 
does occur. However that may be, the difficulty of 
deciding as to the non-existence of tubercles in the 
mammary glands furnishes a sufficient reason for ex- 
cluding from consumption all milk to which such 
suspicion is attached. Such milk is a dangerous food 
for human beings and also constitutes a source of 
infection for animals to which it is fed without pre- 
vious cooking. 

The flesh of cattle affected with tuberculosis and 
slaughtered for the market also becomes virulent un- 
der certain conditions but too little known. A cer- 
tain number of inoculations which have been made 
with muscle juice coming from such animals have 
given. positive results, although in the great majority 
of cases the results of these inoculations have been 
negative. Nevertheless the positive results obtained 
are sufficient to establish the possible danger of the 
flesh of phthisical cattle and to indicate the necessity 



Microbic Diseases Individually Considered. 241 

of compulsory exclusion of such flesh from public 
consumption. This question has been lengthily dis- 
cussed in the various congresses; without insisting 
further on the matter we will say that it is clearly 
connected with the question of the compulsory 
slaughter of all tuberculous animals. Indeed, before 
any diminution in the amount of flesh liable to seizure 
can be obtained the contagion must be' checked be- 
tween the living individuals. But, in the absence of 
any special provision of sanitary police in regard to 
such cases, the owners retain until the last stage, and 
in contact with healthy cows, those animals which, 
on account of their poor condition, they are unable 
to sell but the milk of which still secures for them a 
certain amount of profit. 

We have now reviewed the different means by 
which the germs of tuberculosis are transported from 
diseased to healthy subjects. The receptivity of the 
subject plays an important role in the genesis of the 
process. It is very often dependent upon a special 
predisposition to the disease ; this aptitude to contract 
tuberculosis may be acquired, in which case it results 
from the prolonged influence of bad hygienic condi- 
tions, or it may be transmitted to an individual by his 
ancestors. Heredity of the predisposition is a very 
common occurrence and one especially well recog- 
nized in the human family. 

Acute or chronic catarrhal affections of the respi- 
ratory or alimentary passages favor the implantation 
of the tubercular virus either by diminishing the 
resistance of the tissues and of the organism or by 
producing solutions of continuity by which the germs 
21 



242 Manual of Veterinary Microbiology. 

may obtain entrance. The addition to pulverulent 
virus of bodies of an irregular shape, capable of 
wounding the respiratory mucous membrane, exerts 
a similar influence. Johne has demonstrated the 
great frequency of tuberculosis in cows which respire 
the fumes of iron foundries; these fumes are always 
loaded with fine metallic particles. 

To the number of circumstances favoring the pene- 
tration of the tubercle bacillus into an organism must, 
further, be added the simultaneous presence of other 
microbes which prepare the field for the former. As- 
sociations of this kind may even start into renewed 
activity a tubercular focus which had long been latent 
and regarded as extinct. 

The typical tubercular lesion, the tubercle, is not 
specific of the disease ; it consists in an inflammatory 
nodule of peculiar structure which develops under 
the influence of multiple causes among which may 
be mentioned microbes (actinomyces, zoogloea of 
Malassez and Yignal, pseudo-tubercle bacillus of 
Courmont, etc.), animal and vegetable parasites (de- 
rnodex foliculorum, eggs of the strongylus vasorum, 
aspergillus, etc.), and non-vital agents (powders of can- 
tharides, pepper, and lycopod, and croton oil). The tu- 
bercle bacillus must act by a mechanism similar to the 
preceding causes, that is, through the irritation which 
it excites in the elements which surround it. These 
elements are variable ; they may consist of epithelial 
cells, endothelial cells, or the fixed cells of the con- 
nective tissue, and along with these should be n\en- 
tioned the wandering cells or leucocytes which are 
every-where present and may play an important part. 
The cells in contact with the bacillus increase in size, 



Microbic Diseases Individually Considered. 243 

their nuclei divide, and this is quickly followed by 
the division of the cell body. Simultaneously, the 
elements take a polyhedral form (epithelioid cells). 
This division continues and gives birth to a mass of 
new cells which constitutes the first stage of the 
tubercle. In the course of this initiatory period a 
certain number of elements may break up and dis- 
appear^ but, as a rule, those which are directly in 
contact with the bacillar focus, around the center of 
irritation, acquire considerable dimensions, their nuclei 
continuing to multiply whilst the cell body remains 
single (giant cells) ; these giant cells, however, are 
also formed, at least under certain circumstances, by 
the fusion of several epithelioid cells. When the 
process begins in the interior of the blood vessels the 
leucocytes participate in it from the first, but in the 
opposite case they only intervene at a later stage by 
emigrating toward the invaded parts in order to en- 
ter into the struggle against the intruding germs; 
they then conduct themselves like the elements men- 
tioned above. During this time there is formed at 
the periphery of the epithelioid zone a layer com- 
posed of leucocytes and fixed cells in way of mul- 
tiplication, which in some degree limits or temporarily 
arrests the extension of the process. 

The giant cells especially act as phagocytes ; they 
contend against the bacilli, impair their vitality and 
tend to bring about their degeneration. Metschni- 
koff has described a series of involution forms of 
bacilli in the giant cells of the spermophile. The 
epithelioid and lymphoid cells have the same prop- 
erty. Both giant cells and epithelioid cells may be 
absent when the bacilli are extremely virulent or 



244 Manual of Veterinary Microbiology. 

meet with an organism endowed with great recep- 
tivity; in such case these protective elements have 
not time to become developed and the tubercle is al- 
most entirely lymphoid. 

The extension of the inflammatory process to the 
capillaries involves their obliteration and the absence 
of the reparative plasma in the central parts of the 
tubercle. Thus is explained the necrobiosis (coagu- 
lative necrosis) so frequent in this lesion, although 
the influence of the secretions of the bacillus upon 
the organic elements must also be taken into account 
in this connection. This toxic action of the bacillus 
is the more pronounced in inverse proportion to the 
resisting power of the organism ; thus, in spermo- 
philes the giant cells of which destroy many bacilli and 
establish their superiority over them, caseous masses 
are not found even after a tuberculosis of long dura- 
tion. 

Tubercular lesions are usually localized at the point 
of entry of the germs; the disease is, therefore, pri- 
marily local, and it may definitely remain in this con- 
dition. More frequently, however, it extends, and 
this extension takes place in different ways. In man 
and the large animals extension- occurs chiefly by the 
lymphatics, but it may also occur by way of the blood 
circulation when the bacilli have penetrated into the 
blood through ulceration of a vein at a diseased fo- 
cus, or, indeed, through the medium of the lymphatic 
vessels. 

We have already seen that the first mode of propa- 
gation predominates in the guinea pig, whilst it is 
only accessory, in the rabbit. 

Virulent products coming from one part of the or- 



Microbic Diseases Individually Considered. ' 245 

ganism may be transported to another part by the 
movements of the fluids in certain of the body cavi- 
ties; virulent expectorations which are swallowed by 
the diseased animal can thus infect .the digestive 
canal. , () 

The localization of the lesions is dependent upon 
special predispositions of tissues or organs. This 
fact has been demonstrated by the experiments of 
Schuller, who showed that a contusion produced in 
one of the articulations in a subject artificially in- 
fected by any method whatsoever incites the evolu- 
tion of a tubercular arthritis. 

Tubercular lesions may become purulent in the 
absence of special pyogenic germs; the Koch bacil- 
lus, therefore, secretes a pyogenic substance. The 
injection of a culture in which the bacilli have been 
killed by heat is followed by an abscess. 

Cultures sterilized and freed from bacilli are desti- 
tute of all power of producing tubercles in healthy 
individuals; when injected to tuberculous animals 
they give the reaction of Koch's tuberculin. The 
tubercular poison which occasions the specific neo- 
plasms is, therefore, absent from the soluble portion 
of the cultures; according to Straus and Gamaleia 
it exists in the bacilli themselves and persists after 
their death. Subcutaneous inoculation of dead ba- 
cilli causes a local abscess. Intra-peritoneal inocula- 
tion is followed by a tubercular peritonitis without 
other lesion. Venous injection results in a pulmo- 
nary tuberculosis in which 'the bacilli occur with 
their special staining characteristics unimpaired, but 
the tubercles thus obtained are not infective and have 
no tendency to become generalized. When the dose 



246 Manual of Veterinary Microbiology. 

inoculated is sufficiently large they cause the death 
of the animal like the tubercles of living bacilli, and 
with the same general symptoms. Inoculation of a 
small dose of dead bacilli is followed by temporary 
loss of condition, and the subject becomes much 
more sensitive to a later inoculation with virulent 
bacilli. 

Tuberculosis and scrofula. The lesions of scrofula 
are of a tubercular nature; they contain the Koch 
bacillus. 

M. Arloing has shown that the virus of tuberculo- 
sis and that of scrofula always infect the guinea pig, 
whilst the former only is virulent for the rabbit. He 
has also shown that local tubercular lesions are occa- 
sioned by bacilli which have become more or less at- 
tenuated; sometimes these local lesions are but 
slightly virulent and are of a scrofulous nature; 
sometimes they are more active, resembling tubercu- 
losis properly so called, and like this have a greater 
tendency to extension. 

M. Arloing is of opinion that the bacillus is attenu- 
ated in scrofula; M. Nocard, on the other hand, holds 
that the diminished activity of the scrofulous virus in 
the rabbit is due to the feeble receptivity of this 
species for tuberculosis and to the poverty of this 
virus in bacilli. 

The virulence of scrofulous products is augmented 
by their passage through the organism of the guinea 
pig; whilst tuberculization is slow in the first 
guinea pig it becomes more and more rapid in the 
others inoculated later in the same series. 

These observations concerning scrofula of man are 



Microbic Diseases Individually Considered. 247 

applicable to the scrofulo-tuberculosis of the pig 
(Nocard). 

MM. Courmont and Dor have succeeded in produc- 
ing local articular tuberculosis in the rabbit by intra- 
vascular inoculation of attenuated bacilli ; one or 
several articulations were attacked, whilst the viscera 
remained unaffected. 

Tuberculosis of mammals and avian tuberculosis. 
The question as to the identity of these diseases has 
given rise to much discussion. We will describe first 
of all their differential characters : 

The bacillus of avian tuberculosis is longer than 
that of the disease in mammals. 

It grows on the different media when taken di- 
rectly from the diseased animal, that of the mamma- 
lian disease only, in a satisfactory manner, after 
several transfers upon serum. 

The vegetation of the bacilli of fowls is more rapid ; 
their cultures on solid media are thick, moist and 
luxuriant; those of the bacillus of man are meager, 
dry, scaly and of dull appearance. 

Cultures derived from fowls preserve their vitality 
longer (ten months at least) than those coming from 
man (six months). 

The avian bacillus grows at a temperature as high 
as 43; that of man ceases to grow at 41. 

Avian tuberculosis is with difficulty transmitted to 
the guinea pig and the lesions do not become gener- 
alized as in that which is derived from the human 
being. 

The rabbit, although no more sensitive to the 
tuberculosis of fowls than to that of mammals, shows 
a much great susceptibility to the former than the 



248 Manual of Veterinary Microbiology. 

guinea pig. According to Straus and Gamaleia, avian 
tuberculosis gives rise only to Yersin's septicsemic 
type of the disease when it is inoculated in the veins 
of the rabbit. 

The dog readily contracts" human tuberculosis ; it 
does not take the tuberculosis of fowls. 

Fowls are refractory to human tuberculosis whilst 
very sensitive to inoculations of avian tuberculosis. 
The lesions become localized in the abdominal organs, 
occasionally in the lungs, often also in the marrow of 
bones (causing lameness). 

The question is whether or not these differences are 
sufficiently important to justify us in regarding the 
two bacilli as distinct species. 

Both bacilli have the same form ; the differences in 
size which have been observed are of no importance; 
long bacilli may be seen in mammals and short bacilli 
in fowls. Moreover, both are liable to vary in their 
cultures and according to the animals to which they 
are inoculated; we have seen the bacilli of avian 
tuberculosis become exceptionally short when inocu- 
lated to the calf. 

Both bacilli behave alike toward coloring matters. 
They produce lesions showing the same structure and 
the same general evolution. 

The differential characters drawn from cultures are 
not absolute. 

The inoculation of avian tuberculosis to the guinea 
pig occasionally causes a generalization quite similar 
to that regularly occasioned by human tuberculosis. 
Out of twenty-seven guinea pigs inoculated with the 
spontaneous lesions of the former disease five showed 
a local abscess, seven a discrete visceral tuberculosis, 



Microbic Diseases Individually Considered. 249 

and two the generalized disease (Cadiot, Gilbert and 
Roger). Avian tuberculosis which had acquired 
greater virulence by passing through the organism 
of the rabbit killed seven out of eight guinea pigs 
which received it in the subcutaneous cellular tissue 
(Courmont and Dor). In this case, therefore, the 
bacillus from fowls behaved exactly like that from 
mammals. 

According to an experiment of our own the avian 
bacillus inoculated in the cellular tissue of the calf 
causes a disease exactly like the human bacillus ; as 
with the latter, the evolution is slow and remains for 
a long time limited to the glands receiving the lymph 
from the place of inoculation. 

The non-receptivity of fowls for mammalian tuber- 
culosis is not absolute; out of numerous tests which 
have been made a certain number have given positive 
results ; the lesions, however, were always less gener- 
alized than with the avian tuberculosis. 

Bacilli derived from avian tuberculosis, and which 
had not passed through fowls for at least five years, 
were found to have become more active for mammals 
and produced a generalized tuberculosis in rabbits, 
guinea pigs and chickens. From the liver of one of 
those chickens four guinea pigs were inoculated, none 
of which became tuberculous. A si ngle passage through 
the chicken, therefore, was sufficient to impair the 
virulence for mammals of these avian bacilli which 
previously had acquired a virulence approaching that 
of the human specimen. 

The bacillus of fowls becomes more active for mam 
mals and less active for fowls by passing through the 
organism of mammals. This fact has been proved by 



250 Manual of Veterinary Microbiology. 

MM. Cadiot, Gilbert, and Roger by means of the gen- 
eralized lesions which they obtained in two guinea 
pigs with virus coming from the pheasant. After 
three passages through mammals this virus was inocu- 
lated without effect to two chickens. 

The injection into animals of tuberculin prepared 
from the bacillus of fowls is followed by the same ef- 
fects as the injection of tuberculin of human origin. 

We must therefore regard the bacilli of avian 
tuberculosis and the bacilli of mammalian tubercu- 
losis as varieties of the same species. 

Vaccination. Tuberculosis is essentially a recurrent 
disease; a first attack begets a predisposition to a 
second. It seems, therefore, a priori, paradoxical to 
endeavor to prevent it by means of culture products. 
Nevertheless, MM. Rlchet and Hericourt, Courmont 
and Dor, have succeeded in producing in the rabbit a 
certain degree of immunity against tuberculosis by the 
injection of sterilized cultures of the bacillus of avian 
tuberculosis; they employed bouillon cultures and ob- 
tained sterilization by heat or by filtration. The dis- 
ease was retarded in the majority of the vaccinated 
animals ; in some it was completely prevented. These 
results justify us in affirming the existence of vac- 
cinating substances in cultures of the tubercle bacilli 
and of indulging the hope that, some day, it may be 
possible to vaccinate the human being against this 
terrible disease. 

Vaccination experiments have been made in the 
rabbit by means of the blood serum of the dog 
(hsemocyne). The investigators who have taken the 
initiative in these experiments seem to have obtained 
if not absolute immunity at least a retardation of the 



MicroUc Diseases Individually Considered. 251 

evolution of the experimental disease. The blood of 
dogs previously tuberculized possesses properties more 
active in this respect than that of healthy dogs. These 
results have been utilized in therapeutics, hsemocyne 
having been employed in the treatment of human 
tuberculosis. Tests of the same kind have been made 
with the blood of the goat. 

MicroUc tuberculosis other than that of Koch. We 
have already said that microbes other than the Koch 
bacillus are able to engender the special inflammatory 
reaction which is characteristic of tubercle. Of these 
we are acquainted with several; we will only refer 
specially to the tubercle-begetting zooglcea of Malas- 
sez and Yignal, and to the bacillus of Courmont. 

Zoogloeic tuberculosis of Malassez and Vignal. In 
studying experimental tubercular lesions of the guinea 
pig the authors found, in place of Koch's bacillus, 
micrococci associated in zoogloea. These germs are 
not stained by Ehrlich's method; they maybe demon- 
strated by the following process : 

Sections are left during two or three days in a mix- 
ture of: 

Two per cent solution of carbonate of 
soda, ...... 10 volumes. 

Saturated aniline water, . . 5 " 

Absolute alcohol, . . . 3 " 

Solution made with 9 vols. of distilled 
water and 1 vol. of concentrated so- 
lution of methylene blue in 90 per 
cent alcohol, . . . 3 " 

They are washed with water, dehydrated in abso- 



252 Manual of Veterinary Microbiology. 

lute alcohol colored with methylene blue, and cleared 
in oil of -bergamot or turpentine. 

This solution is a combination of the two solutions, 
staining and decolorizing, of Malassez and Vignal's 
original process (see page. 110). 

The bacterial elements are short, rounded or slightly 
elongated, associated in chains or in small groups, or 
more frequently in large groups or zoogloea. 

Zoogloaic tuberculosis is transmissible from guinea 
pig to guinea pig by inoculation and gives rise to 
generalized lesions like the tuberculosis of Koch; 
but death supervenes at the end of six to ten days, 
that is, much more rapidly than in the case of the 
latter. 

The authors were inclined to think from their first 
observations that the zoogloea and the Koch bacillus 
were the same micro-organism under two different 
forms ; now, however, they must be regarded as ab- 
solutely distinct germs. 

M. Nocard had the opportunity of studying an en- 
zootic of zoogloeic tuberculosis in which all the 
chickens of a farm succumbed to the disease. The 
tubercles, in all cases, had their seat in the lungs, a 
situation in which they are hardly ever seen in gen- 
uine tuberculosis. Moreover, Koch's bacillus was 
absent, whilst the zooglcsse were abundantly repre- 
sented in the lesions. 

MM. Nocard and Masselin produced zooglceic 
tuberculosis by inoculating the guinea pig with the 
nasal discharge coming from a cow suspected of 
phthisis, and which, at the autopsy, was found to be 
exempt from this disease. The cocci found in the 



Microbic Diseases Individually Considered. 253 

guinea pig were cultivated and successfully inocu- 
lated to the guinea pig and the rabbit. 

Bacillar tuberculosis of Courmont. M. Courmont 
found in tubercular lesions of the pleura in an ox 
lesions which did not contain Koch's bacillus a short 
bacillus with its substance condensed at the two ex- 
tremities and with a clear slightly constricted median 
zone; this bacillus is never associated in chains or 
in diplo-bacilli; it is aerobic and anaerobic. It is 
"easily cultivated and grows rapidly in all the culture 
media arid at wide limits of temperature, even up 
to 46. 

Guinea pigs succumb in four to eight days with a 
local oedema and great enlargement of the spleen, but 
without tubercular lesions. The bacilli are abundant 
in the serosity of the oedema and in the blood; after 
several passages through the guinea pig a caseous 
abscess develops at the place of inoculation. 

Rabbits contract a more or less complete tuber- 
culosis; an abscess forms at the place of inoculation, 
and, after death, disseminated or confluent tubercles 
are found in the spleen, liver, and lungs. These 
tubercles have the classic structure ; they do not con- 
tain Koch's bacillus but those described above. 

A culture twenty days old having been inoculated 
to guinea pigs the latter died in less than ten days 
with a generalized tuberculosis in which the lesions 
contained Courmont's bacillus. The property of be- 
getting tubercles in the guinea pig appears to exist 
in cultures only from the twentieth to the twenty- 
fifth day ; at other times the inoculated guinea pig 
dies without tubercjes. The tubercles of the guinea 
pig kill the rabbit, but without tubercular lesions. 



254 Manual of Veterinary Microbiology. 

Courmont's bacillus, therefore, produces a tubercu- 
losis in the ox; the lesions of the ox have direct 
tubercle begetting action on the rabbit but not on the 
guinea pig, except under the special conditions men- 
tioned above. We know, however, that the guinea 
pig is more sensitive to mammalian tuberculosis than 
the rabbit, hence, besides the character of the bacil- 
lus, there is a manifest difference between the two 
diseases. Furthermore, the experimental tubercu- 
losis obtained in the guinea pig by the subcutaneous" 
inoculation of Koch's bacillus develops so slowly 
that, on an average, the lung is invaded only at the 
end of two months; with the bacillus of Courmont, 
as also with the zooglosse of Malassez and Yignal, 
the guinea pig becomes tuberculized in less than ten 
days. Generally, in the rabbit, the duration of Cour- 
mont's tuberculosis is almost the same as that of 
Koch's tuberculosis. Another character important to 
notice is the presence of Courmont's bacillus in the 
blood of subjects tuberculized by means of this bacil- 
lus. Finally, the disease generalizes without infecting 
the lymphatic glands. 

Glanders. 

Glanders is an infectious and contagious disease, 
with progressive course, characterized by circum- 
scribed and multiple alterations localized in the re- 
spiratory mucosa (glanders), or in the skin (farcy). 
These two localizations may coexist, in which case 
we have to do with the disease glanders-farcy. The 
.disease is peculiar to solipeds : horse, ass, and mule ; 
it may also develop in man as a, result of accidental 
inoculation with virulent products coming from one 




Microbic Diseases t Individually Considered. 255 

of these animals ; transmission to the lion, dog, and 
goat has also been observed as a result of spontane- 
ous contamination. 

The disease is acute or chronic ; it is always acute 
in the ass and mule, as well as in the lion.* 

Microbe. The bacillus mallei is a Blender, motile 
rod, straight or slightly curved, and with rounded 
ends. It measures 2/j. to 5// in 
length by 0-5// to 1-5/* in thick- 
ness, hence it is thicker than the 
Koch bacillus, but of the same 
length. When stained it shows 
alternating clear and colored 
spaces which give it a granular 

aspect recalling that of the tu- , n ^ ci11 " 8 mallei ; X 
u i i, MI T> ^ i 1 00a From a photo- 

bercle bacillus. Eosenthal re- micrograph . (F rankel 

gards these clear spaces as spores ; and Pfeifer.) From 

others, however, basing them- Sternberg's Bacteriol- 

selves on the slight resistance of 

the bacillus to heat, refuse to admit the existence of 

spores.(l) 

The glanders bacillus is aerobic ; it occurs in the 
pathological secretions nasal discharge, pus of ul- 
cers ; and in the specific lesions farcy buds, tuber- 
cles, glanderous ulcers, and inflammation of the cor- 
responding lymphatic vessels and glands, etc. It is 

(1) Bab^s has shown that such granules are present in the ma- 
jority of bacilli, even the least resistant, and that they do not 
possess the resisting power of true spores. He believes that they 
exercise the same function in the multiplication of bacilli as the 
chromatic part of cell-nuclei in the division of cells. 

* [The sub-acute or chronic form ol the disease seems to be not 
uncommon in mules in the Southern States. D.j 



256 Manual of Veterinary Microbiology. 

I present in the blood only in acute forms, and then in 

1 very small numbers. 

Action of physical and chemical agents. The con- 
tagion of glanders is destroyed by two minutes ex- 
posure to a temperature of 100, but its destruction 
can be obtained at lower temperature, provided the 
action of the heat is more prolonged, for example, 
five minutes at 65, ten minutes at 55. Glanderous 
pus, spread in a thin layer and left to desiccate in 
contact with the air, loses its activity between the 
second and third days; hot and dry weather fa- 
vors its destruction, while cold and wet weather 
retards it. Under the same conditions of desic- 
cation, but excluded from the air, it yet shows 
itself active after twenty-six days. Virus rapidly 
and thoroughly dried retains its vitality in contact 
with the air longer than that which is slowly and im- 
perfectly dried. The discharge from the nose in gland- 
ers, when immersed in water, has been found to re- 
tain its virulence for eighteen days. Virulence is not 
readily destroyed by putrefaction; inoculations made 
with the central part of pieces of glanderous lungs 
abandoned to the air for fifteen, eighteen, and even 
twenty-six days, have given positive results. (Cadeac 
and Malet.) 

The following substances destroy the virus of 
glanders after one hour of contact : carbolic acid, 2 
per cent; sulphuric acid, 2 per cent ; chloride of zinc, 
2 per cent; saturated lime water; hypochlorite of 
lime, 1 per cent; corrosive sublimate, 1 to 1,000, 
and 1 to 10,000 ; sulfate of copper, 5 per cent ; per- 
manganate of potash, 5 per cent ; nitrate of silver, 1 



Microbic Diseases Individually Considered. 257 

to 1,000; chlorine gas, and sulphurous acid gas, in 
concentrated solution. 

Cultures. Cultures are easily obtained on the dif- 
ferent media; they require free access of atmos- 
pheric oxygen ; 37 is the most favorable tempera- 
ture; below 20 their growth is arrested except upon 
glycerin-agar ; it also ceases at 43, and the germs 
are killed at 55. 

Bouillons become turbid within twenty four hours 
without presenting special characters. 

On agar and on serum it shows a bluish- white 
translucid growth in the form of droplets, or in a 
continuous layer which becomes opaque as it increases 
in thickness. 

Potato is admirably adapted to the culture of the ba- 
cillus of glanders. On this medium it forms a thick, 
moist, glistening, viscid coating, which after a few days 
assumes a fawn color, gradually deepening to a bright 
chocolate. (Nocard.) This culture is characteristic 
and should assist us in the diagnosis of doubtful 
cases; it suffices to sow a particle of the suspected 
product, previously diluted, upon potato ; the latter 
soon becomes covered with diverse growths among 
which we should recognize by their peculiar color 
those due to the glanders bacillus.* 

Research and coloration. The bacillus mallei shows 
little affinity for the aniline colors; it does not 
support the Gram or Weigert stains. The methods 

* [Such simple culture tests can be of value when the suspected 
material is obtained from a yet unopened skin nodule or from an 
extirpated submaxillary lymph node, but will rarely assist us in 
the examination of nasal discharge. D.] 
22 



258 Manual of Veterinary Microbiology. 

of double staining are therefore inapplicable. Gen- 
erally we have recourse to Loffler's blue, decolorizing 
with carbolated water, at 1 to 300. 

Kiihne also recommends the following method of 
staining: Sections, well freed from alcohol, or cover 
glasses, are placed in a solution composed of: water, 
100; carbolic acid, 5; alcohol, 10; methyl ene blue, 
1-5. They are decolorized by a rapid passage 
through water acidulated with hydrochloric acid, and 
washed in distilled water. The sections are dehy- 
drated by a short immersion in alcohol, then placed 
in aniline oil to which has been added a few drops of 
oil of turpentine, then in pure turpentine, and, 
finally, in xylol. 

The bacillus of glanders is difficult to demonstrate 
in old lesions, where it appears to break up into 
granules (perhaps into spores); it is always much 
more abundant in acute lesions. 

Experimental inoculations. Solipeds, the sheep, goat, 
pig, dog, cat, rabbit, guinea pig, field mouse and pig- 
eon take the disease by inoculation. The ox, white 
mouse, rat and chicken are refractory. The ass 
always contracts acute glanders by inoculation. 
When the virus is inserted by scarifications in the 
forehead, for example, an extensive ulcer, with indu- 
rated and inflamed borders, develops on this region. 
Along with this the manifestations of the general 
disease appear about the third day, and the subject 
quickly dies. The autopsy reveals an eruption of 
small reddish nodules on the respiratory mucous 
membrane, pysemic infarcts in the lungs, liver, kid- 
neys, spleen, marrow of bones, etc. 

In the dog the insertion of the virus in the skin or 



Microbic Diseases Individually Considered. 259 

the subcutaneous cellular tissue is sometimes followed 
by a violent access of fever in the course of the third 
day ; then the local lesions appear in the form of an 
extensive engorgement which bursts and ulcerates, 
the chancre thus produced having a great tendency 
to spread. Farcy buds and cords, and rodent ulcers 
often appear on other parts of the body; sometimes 
an intense and painful lameness expression of glan- 
derous arthritis supervenes without apparent cause, 
the stifle being the most frequent seat of the specific 
inflammations. Out of twelve dogs inoculated by 
Prof. Reul, four died, three were killed, and five 
spontaneously recovered. Inoculation of glanders to 
the dog often gives, as sole reaction, a superficial 
ulceration which cicatrizes in eight to fifteen days. 

In the guinea pig inoculation by scarifications 
(these are made on the neck or back) is followed by 
ulceration of the wounds on the fifth to the tenth 
day; these ulcers may, later, cicatrize. Subcuta- 
neous inoculation gives voluminous abscesses in the 
whole chain of lymphatic glands proximal to the 
place of inoculation. In both cases the animal be- 
comes emaciated and dies more or less rapidly with 
tubercular foci in the spleen, liver, lung and lymphatic 
glands. The local lesion the ulcer suffices to es- 
tablish the glanderous nature of the inoculated prod- 
ucts. "When the inoculation is made in the perito- 
neum a pronounced swelling of the scrotum is ob- 
served from the second to the third day, this swelling 
indicating the specific inflammation of the testicular 
membranes. This peculiarity can be utilized for the 
purpose of making a rapid diagnosis of the disease 
(Straus). 



260 Manual of Veterinary Microbiology. 

When glanders is inoculated to the dog or guinea 
pig by scarifications it may be followed by a negative 
result. Hence, an absolute value should be assigned 
only to positive results, and the suspected material 
which has been unsuccessfully inoculated submitted 
to further tests. 

Diagnosis of doubtful cases. Besides experimental 
inoculations to susceptible animals, guinea pigs, dog, 
and ass, and the cultures on potato of the suspected 
discharge, some have counseled the employment of 
auto-inoculation. This operation consists in inocu- 
lating a horse with the products, supposed to be viru- 
lent, coming from the same animal ; these insertions are 
made in the skin by puncture or superficial incision and 
the existence of glanders confirmed if the operation is 
followed by the formation of a chancre; but, as we 
shall see later, no conclusion can be reached founded 
on the absence of the ulcerous reaction. In default 
of any discharge some have resorted to extraction of t 
a swollen lymph node and examination of the same 
for the specific bacilli. Finally, we may have re- 
course to test inoculations with mallein. 

Mallein. By this name is designated a glycerin 
extract of pure cultures of the bacillus of glanders. 
This is the glanders lymph, the analogue of the 
tuberculosis lymph or tuberculin. The extract is 
sterilized by heat, and diluted to ten times its weight 
with two per cent carbolic solution. 

The injection of SO to 50 centigrams of this 
dilution to glanderous horses produces a reaction 
characterized especially by dejection, acceleration of 
the pulse, and elevation of temperature ranging from 
1*5 to 2, by a hot redematous swelling as large as 



Microbic Diseases Individually Considered. 261 

the hand at the place of inoculation, and by an ap- 
preciable swelling of the subglossal lymphatic glands. 
In a healthy animal the same injection would produce 
no effect. Mallein should, therefore, facilitate the 
diagnosis of the disease in doubtful cases. 

Etiology and pathogeny. The efficient cause of 
glanders is the bacillus maRei. This microbe, under 
natural conditions, multiplies only in the organism 
of solipeds and ita origin must be looked for in 
these animals. The virulent substances cast off by a 
glanderous subject are : nasal discharge, pus of ulcers, 
saliva, urine, pus of setons, and semen ; the virulence 
of the last four products, though less constant than 
that of the first, is nevertheless incontestable and has 
been demonstrated by experiment. To this list must 
be added, in the case of the cadaver, the various 
specific lesions and the muscles. Inoculation of 
guinea pigs with muscle juice has produced the dis- 
ease in a certain number of cases. 

Contagion takes place by direct or indirect con- 
tact. There is direct contact from the horse to man 
when the latter inoculates himself in manipulating 
or in dressing glanderous lesions; from horse to 
horse when two horses, one of which is glandered, 
occupy adjacent stalls in a stable, or work side by 
side so that they can easily touch each other. The 
disease is occasionally transmitted through sexual in- 
tercourse ; Zundel mentions the case of a glandered 
stallion which infected more than fifty mares. The 
eventual passage of the glanders bacillus into the 
semen accounts for this mode of infection, whilst 
their filtration through the placenta occasions trans- 
mission of the disease from mother to foetus, an 



262 Manual of Veterinary Microbiology. 

occurrence which has been observed by MM. Cadeac 
and Malet. Inter-sexual and intra-uterine contami- 
nation are, however, actually of rare occurrence. 

Indirect contagion is much the most frequent. The 
glanders virus, distributed externally, contaminates 
the food, drinking water and litter, harness, groom- 
ing utensils, sponges, brushes, curry-combs, etc., the 
walls of houses, mangers, racks, and such like; if 
it meets with the conditions necessary for desicca- 
tion it will be carried with the dust into the sur- 
rounding atmosphere. The morbific germ is trans- 
ported on to the healthy organism through the in- 
termediation of numerous vehicles. Infection may 
take place through the respiratory passages, the ani- 
mal inspiring air charged with virulent particles, but 
this mode of contagion should be rarely effective in 
glanders since desiccation is a puissant cause of the 
destruction of the bacillus. The disease is more cer- 
tainly communicated by the digestive canal, through 
swallowing infected food or water. This contamina- 
tion of the ingesta may result from their immediate 
contact with the products coming from a diseased 
animal or it may occur during their storage in in- 
fected places. 

In this connection should be mentioned the danger 
arising from the consumption of animals which have 
died or been killed while suffering from -the disease. 
This danger exists not only for the specific lesions, 
in which the virulence ts evident, but also for the 
flesh. The investigations of MM. Cadeac and Malet 
have shown that the juice of such flesh is capable of 
communicating glanders. 

Finally, the virus may penetrate through the skin 



Microbic Diseases Individually Considered. 263 

to which it has been carried in various ways and es- 
pecially by the harness, grooming utensils, litter, etc.; 
it gains entrance through accidental abrasions of the 
integument. According to Babes, however, the 
glanders virus can make its way into the organism 
through the intact skin, penetrating the orifices of 
the hair follicles. This penetration will by facilitated 
by frictions. M. Nocard undertook to test this asser- 
tion by rubbing an ointment charged with glanders 
bacilli on the skin of three asses and fifteen guinea 
pigs. Of these, only two guinea pigs became gland- 
ered, a result which considerably reduces the risk 
which might be inferred from the above conclusions. 

Certain circumstances are of such a nature as to 
favor the implantation of the bacilli of glanders. 
We recognize the predisposing influence of bad hy- 
gienic conditions, excessive fatigue, and chronic ex- 
hausting diseases. Chronic forms of glanders may 
assume acute characters under the same influences. 

The bacilli of glanders having once obtained en- 
trance into the economy multiply generally at the place 
of their penetration, thus producing local lesions. 
They quickly invade the lypmphatic system (which 
becomes the seat of specific inflammations, glander- 
ous lymphangitis and adenitis) and the blood; by the 
latter they are carried throughout the system but 
they only develop in the tissues predisposed to their 
attack: respiratory mircous membrane, integument, 
testicles, and synovial membrane of articulations and 
tendons, etc. 

The specific lesions present certain analogies with 
those of tuberculosis. The primary lesion the glan- 
ders tubercle is purulent at its center and destitute 



264 Manual of Veterinary Microbiology. 

of giant cells; besides this marked tendency to sup- 
puration, should be mentioned the early retrogressions 
which lead to the ulcerations characteristic of the dis- 
ease. 

Vaccinations. Glanders leaves behind it no immu- 
nity against another attack. Thus, auto-inoculations 
and re-inoculations performed upon animals already 
affected with the disease are followed by positive re- 
sults. In the glanderous guinea pig these inoculations 
determine local symptoms as intense and a generali- 
zation as complete as at the time of a first insertion; 
in the dog the second attack is also generalized and 
often as severe as the first. According to Galtier the 
dog can contract the disease as often as five times, 
but the local symptoms become less and less marked 
In the horse re-inoculations and auto-inoculations 
may prove abortive or may produce a chancre with 
or without lesions of the lymphatics, but fever and ag- 
gravation of symptoms are never observed, the virus 
seeming to limit its effects to the point of inocula- 
tion. In the ass secondary inoculations are followed 
by much more intense reactions; each insertion pro- 
duces a marked tumefaction and a corresponding 
ulcer, corded lymphatics, glandular engorgements 
in brief the effects of a first inoculation. (Cadeac 
and Malet.) 

The natural disease, therefore, does not confer im- 
munity. Several attempts at artificial vaccination 
have been made. Straus obtained immunity in a dog 
l>y the intravenous injection of a small quantity of 
virulent culture. A bei>ign disease is produced which 
immunises the dog against intravenous inoculation of 
large doses. 



Microbic Diseases Individually Considered. 265 

Epizootic lymphangitis, or African farcy. 

Hivolta has described, and M. Nocard confirmed, 
the presence in the pus and lesions of African farcy 
of " a sort of micrococcus, slightly ovoid and some- 
what pointed at one of its extremities, measuring 3// 
to 4jj, in diameter; its contour is clearly defined by a 
very refringent line." This organism (cryptococcus 
of Bivolta) is colored by the Gram-Weigert-Kiihne 
methods ; but its dimensions and its refringence are 
such that it is impossible, according to M. !N~ocard, to 
confound it, even when unstained, with any other 
element. 

Several practitioners have described the appearance 
of chancres of acute glanders on the nasal mucosa of 
animals attacked by epizootic lymphangitis. The 
demonstration of the cryptococcus in these lesions 
enabled M. Nocard to affirm that they were related to 
lymphangitis and not to glanders. Moreover, the 
bacillus of this last disease was lacking. 

Strangles.* 

We are indebted to Schiitz for the investigation of 
the microbe of strangles. It is a streptococcus which 
occurs in short chains, in diplo- and in monococci in 
the nasal discharge and in the pus of the lymph- 
glandular inflammations and abscesses symptomatic 
of the disease; in long, tortuous chaplets in sections 
of the inflamed organs. It readily takes the differ- 
ent aniline stains. Inoculated under the skin of the 

* [Also called, in America, " Distemper " of horses ; Fr.. 
Gourine ; Ger. Druse. D.] 
23 



266 Manual of Veterinary Microbiology. 



Fig. 13. 



horse it causes the formation of an abscess. In the 
mouse* it produces, in addition, metastases by way 
of the lymphatics and blood vessels. Natural in- 
fection takes place through different channels but 
more especially through the respiratory and digestive 

mucous mem- 
branes. Its ab- 
sorption is facili- 
tated by the 
presence of solu- 
tions of continu- 
ity, but it has 

not ^ een snown 
that these are 

necessary. Ac- 
cidental or ope- 
rative wounds 
also sometimes 
furnish ports of 
Streptococcus equi, pus of strangles of horse, entry for the 
XlOOO. (Kitt.) virus; castration 

wounds, for example, may be infected by the surgeon 
himself, his hands or instruments being soiled with 
the germs from previously handling horses suffering 
from strangles. The disease is transmitted from the 
mother to the foetus. Whatever may be its mode of 

* [An important differential test in distinguishing this disease 
(in its less characteristic forms) from glanders consists in the in- 
oculation of mice. White and gray house mice are highly sus- 
ceptible and die from inoculation with strangles, but are insus- 
ceptible to glanders. Field mice, on the contrary, die from inocu- 
lation with glanders, but develop only a local lesion from inocula- 
tion with strangles. Th. Kitt : Bacterienkunde, p. 254. D.] 




Microbic Diseases Individually Considered, 267 

entry the germ appears to pass at once into the blood 
as is indicated by the high fever at the beginning of 
the disease; it then becomes localized in the lym- 
phatic system. The microbic theory of strangles ac- 
counts for its contagiousness, the secondary abscesses 
met with in the course of the disease, and the sep- 
tico-pygemic complications which, in certain cases, 
terminate it. 

Contagious acne of the horse. 

This disease is characterized by a pustulous der- 
matitis which is very readily transmitted to other 
animals. It is generally benign hut may be more 
severe and lead to the production of ulcerations and 
inflammations of the lymphatic vessels and glands. 
It is, however, always easily distinguished from farcy 
by the great tendency to cicatrization of the ulcers 
which it occasions ; in farcy, moreover, true pustules 
do not occur. Acne is distinguished from horse-pox 
or variola of the horse by the fact that the eruption 
in this last disease is always localized in the lips, 
nostrils, and pasterns. 

Dieckerhoff and Grawitz discovered in the pus of 
acne a short bacillus (2//) which they cultivated and 
the culture of which, rubbed into the skin of the 
horse, reproduced the disease. The guinea pig suc- 
cumbs in twenty-four hours when subjected to the 
same treatment. These cultures are also pathogenic 
for the ox, sheep, dog, and rabbit. Inoculated under 
the skin of the mouse it gives rise to the formation of 
an abscess. 

The natural disease is transmitted by means of the 
grooming utensils, harness, blankets, etc.; thus, it is 



268 Manual of Veterinary Microbiology. 

not uncommon to see it localized on the regions cov- 
ered by the saddle or girth. 

Actinomycosis. 

Actinomycosis is a disease, most frequently of a 
local character, caused by a vegetable parasite, the 
actinomyces. The lesions by which it is character- 
ized have long been recognized in practice, but their 
true nature was entirely unknown before the inves- 
tigations of Bellinger, the different names, such as 
osteosarcoma, osteoporosis, spina-ventosa, cancer or farcy 
of the bones, etc., by which the maxillary tumors of 
cattle were designated reflecting the very diverse and 
somewhat vague opinions of the various writers upon 
these productions. 

The discovery of a special fungus in the majority 
of these tumors has enabled us to classify them with 
the parasitic infections and explain their great in- 
tractability.. 

The facts established by the authors in their first 
communication on actinomycosis have been repeat- 
edly confirmed since then in both human and vet- 
erinary medicine, and, referring here to the latter 
branch of medicine only, the lesions of this disease 
have been found in other parts of the body than in 
the maxillae. They have been met with in the 
tongue, pharynx, reticulum, liver, nasal cavities, 
larynx, lungs, neck, and vertebrae. 

In the pig they have been observed in the muscles, 
lungs, amygdalae and mammae; in the horse in cer- 
tain cases of scirrhous cord. A case has also been re- 
corded in the dog. 

Characters of the parasite. The actinomyces 



Microbic Diseases Individually Considered. 269 

fungus) occurs in the specific tumors in the form of 
yellow or occasionally colorless grains ; when the ma- 
terial in which they are contained is spread out on a 
glass lip the smallest of these grains appears some- 
what like grains of sand ; the larger are formed by 
the union of primary granules and are of various 
forms. These grains are most frequently cretaceous, 
having a hard, stony consistence. In size they vary 
from 0*1 millimeter to 1 millimeter, or more. 

They are composed of one or several colonies of a 
fungus the elements of which are arranged in rays. 
In each colony there may be distinguished : 

1st. A central zone, formed of very fine filaments 
ramifying and intermingled in a close felted net-work. 
The diameter of these filaments is uniform in all parts 
of the central zone; in structure they consist of hol- 
low cylinders, each, at intervals, containing a nucleus 
which readily takes up coloring matters. (1) 

The size of this central part of the actinomyces 
corresponds with that of the tuft or colony; small in 
microscopic grains it is of much larger dimensions 
in those tufts which have acquired a considerable 
volume. 

2d. A peripheral zone, rendered conspicuous by the 
radiating elements of which it is composed ; these 
elements are pyriform with their large extremities 
losing themselves in the tissue which surrounds the 
colony and even occasionally penetrating into the 
adjacent cells, and their slender ends passing into the 

(1) Actinomycosis and its parasite. See Annales de medicine 
veterinaire, 1890. 



270 Manual of Veterinary Microbiology. 

central zone with the filaments of which they become 
continuous. 

The thickness of the peripheral zone is quite uni- 
form whatever may be the size of the actinomyces 
itself, but it may be unequal in the different parts of 
the same colony on account of the varying dimensions 
of the cortical enlargements ; the average length of 
these enlargements is from 15// to 30/z and breadth 
5/* to 7//, but these limits may be much surpassed; 
we have seen some which measured 74/* in length by 
10// in thickness. 

The club-shaped enlargements may be simple or 
branching, and branching may take place either from 
the slender pedicle or from the enlarged part itself; 
the branches, emerging in this manner, may them- 
selves divide giving rise to new club-shaped swell- 
ings, so that the whole obtains a more or less dense 
arborescent appearance. These club-shaped enlarge- 
ments are composed of a resistant membrane and 
clear contents ; occasionally the membrane shows 
circular depressions which seem to divide the body 
into small cubical elements, and at these points trans- 
verse division is readily produced. 

Besides the typical actinomyces just described 
there are others which are quite small and in which 
the enlargements are absent, and others, again, in 
which the filamentous central zone seems to be en- 
tirely transformed into these club-shaped elements. 

In colonies of large size the central zone contains 
micrococci, appearing like small round points of less 
than 1/z in diameter, united in chains, or, more fre- 
quently, in small irregular masses ; these nests of 
micrococci, in the largest tufts, are not infrequently 



.Microbic Diseases Individually Considered. 271 

found in large numbers, their nature being more 
easily recognized by the rarefaction of the mycelial 
felt-work in these situations. 

Fig. 14. 




Tuft of Actinomyces. Isolated clubs of Actino- 

(From Kitt's Bacterienkunde.) myces. (Johne.) Do. 

MM. Cornil and Babes have described a special 
condition of the filaments of the periphery of the 
colonies, in which they terminated by various slight 
enlargements bearing conidia. 

Harz classes the actinomyces with the hyphomy- 
cetes fun^jpi regarding it as a complete fungus com- 
posed of mycelia, hyphse and spores, the mycelium be- 
ing represented by a basal cell from which spring the 
hyphce the branching filaments of the internal zone 
and these bearing the spores or enlargements at the 
periphery. 

The basal cell of Harz has not been found by other 
investigators; most of these, however, agree in re- 
garding the enlargements as spores or, rather, as 
sporangia ; hence these enlargements have received 
the name of conidia. 

The classification of the actinomyces with the fungi 
appears to us to be supported only by the results of the 



272 Manual of Veterinary Microbiology. 

examination of this parasite in the tumors to which 
it gives rise. 

Bostroem classes it with the group of schizomy- 
cetes and especially with the cladothrix, a conclusion 
which is forced upon us from a study of the parasite 
in its cultures, and one to -which our own researches 
have led us. We are also inclined to regard the mi- 
crococci, spoken of above, as spores. 

The actinomyces is anaerobic and facultatively 
aerobic. 

Action of physical and chemical agents. According 
to Domec, the filaments are killed in five minutes by 
moist heat at 60, whilst the spores resist for that 
length of time a temperature above 60 but below 
75. This feeble resistance of the spores compared 
with that of the spores of bacteria has been advanced 
as an argument in favor of regarding these spores as 
allied to those of the mucedinese, and of classifying 
the parasite with this group of plants. 

The action of chemical agents has been little 
studied. Iodine has been recommendedtfor the de- 
struction of the actinomyces in the tumors, but the 
hot iron is still the means most to be recommended 
for this purpose when it is possible to use it. 

Research and coloration. The examination of the 
actinomyces fungus is extremely simple, the suspected 
material merely requiring to be spread upon a glass 
slide; the hard, yellow grains which to the naked 
eye have the appearance of grains of sand, separate 
themselves during the operation. These grains are 
the colonies of the actinomyces. To verify this it is 
merely necessary to put on a cover glass and examine 
with the microscope ; a magnification of from two to 



Microbic Diseases Individually Considered. 273 

three hundred diameters will always prove sufficient 
for the recognition of the parasites. 

This mode of preparaton, however, is inadequate 
for the detailed study of the fungus. The center of 
the radiating masses is nearly always calcified, and 
this transformation conceals its filamentary structure 
and, at the same time, prevents the dissociation of the 
tuft. The latter is also enveloped by a mucoid, viscid 
substance which tends to hold together the enlarge- 
ments at the periphery. The inconvenience arising 
from these two circumstances is obviated by treating 
the substance to be examined with alkalies or dilute 
acids. 

For this purpose we have found the employment 
of dilute ammonia quite satisfactory ; the sand-like 
grains yet require to be crushed in order to spread, or 
better, to dissociate them. 

By this means beautiful preparations are obtained 
in which the mycelium and its connection with the 
conidia can be studied. 

The examination of sections is highly instructive. 
The actinomyces can be stained in different ways. 
According to the way which is mostly recommended, 
the sections are immersed in a solution of orseille,(l) 
then in alcohol, and, finally, in an aqueous solution 
of gentian violet or methylene blue. A double stain- 
ing is thus obtained, the conidia being red and the 
mycelium violet or blue. 

In our opinion Weigert's method gives better re- 
sults. The antinomyces take a violet color and the 

(1) Pure orseille, free from ammonia, dissolved until a deep red 
color is produced, in : acetic acid, 5; absolute alcohol, 20; water, 40, 



274 Manual of Veterinary Microbiology. 

tissues of the tumor may be stained red by picro- 
carmine. 

The methyl violet, by this method, may act in two 
degrees. When the preparation has been exposed to 
the staining solution for a short time the mycelium 
of the actinomyces is alone colored violet, when it has 
been longer exposed mycelium and conidia are stained 
alike. 

Preparations of some value may also be obtained 
by the employment of picro-carmine alone ; the fungi 
are stained yellow, whilst the neoplasm is red, but 
by this process we do not obtain a definition of the 
structure of the parasite. 

MM. Cornil and Babes recommend double staining 
by the method of Gram with eosin or safranin. 
They thus obtain a violet staining of the central fila- 
ments whilst the clubs or conidia are stained red. 

Cultures. Bostroem obtained cultures upon beef 
blood-serum and upon agar, and has been able .to 
convince himself that the bright claviform enlarge- 
ments of the periphery of the colonies are incapable 
of multiplication, this property belonging to the cen- 
tral filaments alone ; hence, he proposed classing the 
actinomyces with the schizomycetes or bacteria. The 
structure would be that of a cladothrix or branching 
bacterium, and the swollen productions of the corti- 
cal zone only incidental forms developing when the 
parasite finds itself under special conditions of nutri- 
tion. 

"We have cultivated the actinomyces in alkaline 
bouillons and our researches have led us to the same 
conclusions. Inoculation of peptonized glycerin- 
bouillon with a particle of tumor produces a devel- 



Microbic Diseases Individually Considered. 275 

opment already well marked at the end of the first 
day; at this time may be seen a viscid, coherent layer 
floating near the bottom of the nutrient fluid which 
is still perfectly limpid, although holding in suspen- 
sion a few white or yellowish, irregularly shaped 
grains. In examining these last under the micro- 
scope, they are found to be composed of ramified 
filaments, a few of which bear one or more distinct 
enlargements in every respect similar to the conidia 
of the cortical zone of the actinomyces. This struc- 
ture is very evident, the mycelial filaments being yet 
loosely intermingled. Each of the latter consists of 
a tube containing, from point to point, a rounded 
nucleus which has a well marked affinity for the 
aniline colors. The bouillon acquires a fetid odor. 

On the following days the pullulation continues 
with great activity, but the culture changes its ap- 
pearance after the third day; the slimy growth in the 
bulbs resolves itself into a cloud of very fine granules 
which fall to the bottom of the bouillon as if the mu- 
coid substance which held them suspended had 
disappeared; at the same time, the two layers of the 
liquid, up till now clearly defined, spontaneously mix 
themselves. 

In examining bouillon cultures with the micro- 
scope, after the third day, the claviform enlargements 
are no longer to be seen. The small white granules 
deposited at the bottom of the bulbs are found to be 
composed solely of fine filaments, presenting lateral 
ramifications more abundantly than in the preceding 
preparations, and showing always in their interior 
the rounded nuclei, placed at regular intervals; 
these mycelial tubes are united in felted balls from 



276 Manual of Veterinary Microbiology. 

which some filaments separate themselves to extend 
outward, or they form small tufts the branches of 
which radiate around a certain point in one of these 
filaments. Besides these groups of filaments a few 
may be observed .which seem detached, but which 
have the same structure and are also branched. 

We have succeeded in obtaining several genera- 
tions of these cultures, starting from a tumor of the 
ox. We have also made cultures with the products 
obtained by inoculating the disease to rabbits. In 
these last cases the results have been the same, ex- 
cept that we have never seen any thing resembling 
the conidia. 

Cultures are much more active when protected 
from the air; the actinomyces is, therefore, chiefly 
anaerobic. Various investigators have cultivated the 
fungus upon potato ; the surface of the latter be- 
comes excavated during the first days, then covered 
with colorless colonies with irregular surface, which 
soon becomes prominent and powdery ; they become 
gray in color, then yellow, or even greenish when 
exposed to the light. Domec, in studying these cul- 
tures, has recently come to the conclusion that the 
actinomyces is a mucedine. 

Experimental inoculations. Johne has transmitted 
the disease to the calf and to the cow by subcuta- 
neous, intra-peritoneal, and mammary inoculation; 
Ponfick and Israel have communicated it to the 
calf and to the rabbit. We, also, have inoculated 
actinomycosis to the rabbit both with artificial cul- 
tures and with the natural products obtained from a 
cow. The lesions remained local in all cases, and 
consisted of an exhausting suppuration which in- 



Microbic Diseases Individually Considered. 277 

volved the death of the subject in from seven to ten 
days. 

Etiology and pathogeny. The aetinomyces infects 
only the herbivora and omnivora ; hence, the dis- 
ease has been attributed to the forage. Johne found 
an identical fungus on the surface of husks of barley 
arrested in the tonsils of a healthy pig. Piana dis- 
covered, in a tumor of the tongue, vegetable debris 
along with the actinomyces. Most writers agree in 
incriminating more especially straw and barley husks. 

The parasites, distributed on the ground by diseased 
animals readily bring about the transmission of the 
disease to healthy animals ; Stienon records an in- 
stance of enzootic actinomycosis in which nearly all 
the cattle of the farm were affected. 

Inoculation has also been successfully performed by 
Johne. This author injected under the skin and 
within the abdomen of two calves, and into the ud- 
der of a cow, the juice of a tumor of the maxilla and 
by this means produced the characteristic neoplasms. 
Similar results have been obtained by Ponfick and 
Israel in the calf and in the rabbit. This last author 
made his inoculations with the actinomycosis of man ; 
the effects obtained were the same as with the tumors 
of cattle. Finally, accidental contamination has 
been established in persons who have attended to 
animals affected with the disease. 

The lesions being most frequently confined to some 
portion of the digestive canal, it is logical to infer 
that the natural infection occurs through accidental 
solutions of continuity of the mucosa of these pas- 
sages, principally in the anterior passages (wounds of 
the gums, carious teeth, crypts of the tonsils, etc.) 



278 Manual of Veterinary Microbiology. 

But penetration may also probably take place through 
the respiratory tract, the germs being transported 
by the air, by foreign bodies, or merely by the mu- 
cus. 

Cutaneous wounds are also channels of introduc- 
tion of the parasite (actinomycosis of the testicular 
cord in the horse ; actinomycosis of the leg in the 
same animal in consequence of a kick ; subcutaneous 
abscesses in cattle). 

The tumors of actinomycosis (actinomycomata) re- 
sult from the irritation excited by the actinomyces ; 
they frequently have the structure of granulomata ; 
the parasite nearly always occupies the center of an 
inflammatory nodule which may readily be differen- 
tiajed into two zones, an internal, formedof epithelioid 
cells, and a peripheral, containing fusiform and lymph- 
oid cells and indicating the transition to the con- 
densed connective tissue which forms the frame work 
of the tumor. The fungus may also very frequently 
be seen surrounded by a row of giant cells. This 
structure resembles that of the tubercle type and 
shows that the pathogenic action of the actinomyces 
consists in a circumscribed and chronic irritation of 
the tissues into which it has been carried. After 
these tumors become softened in the center they con- 
tain a puriform yellowish or white liquid in which 
are suspended yellow grains, most frequently hard 
and calcified; these grains are the actinomyces. 
When the softening is regular the tumor acquires a 
characteristic spongy appearance. 

It may happen, and this is the rule in man, that 
the actinomycotic tumors become complicated with 
suppuration. It appears to be established that the 



Microbie Diseases Individually Considered. 279 

formation of pus should be attributed to tbe acti- 
nomyees themselves and not to pyogenic bacteria as- 
sociated with them. Without excluding in all cases 
the intervention of the latter we think that the pres- 
ence or absence of suppuration may be dependent 
upon the animal species infected, as well as on the 
tissue invaded. Thus, the injection of pure cultures 
of actinomyces within the abdomen of the rabbit 
always occasions a suppurative peritonitis ; in man, 
also, the tendency to suppuration is always very 
marked. Stienon records an instance of cattle af- 
fected with subcutaneous abscesses of an actinomy- 
cotic nature. 

The tissue of the organ in which the tumor de- 
velops shows alterations corresponding to the chronic 
inflammations set up by its presence ; certain parts 
become hypertrophied, whilst others diminish or may 
even disappear. Thus are explained the fibrous in- 
duration of some of these tumors, the osseous stalac- 
tites which develop on the maxilla when the invasion 
begins in the periosteum, and the cavities into which 
the same bone becomes hollowed when the path- 
ological process takes place in its center. 

Lesions may appear in the neighborhood of the 
first affected part ; they result from the progressive 
enlargement of the primary tumor, or from a vascu- 
lar emigration of the fungus. 

The gradual increase in size of the neoplasm may 
lead to its extension to the skin or may give rise to 
distension, thinning, arid perforation of the latter in 
one or more points, from which protrude fungus-like 
growths which ulcerate and secrete p'us mixed with 
the yellow, pathognonionic grains. 



280 Manual of Veterinary Microbiology. 

The lymphatic vessels may convey the parasite to 
the corresponding glands and the latter then become 
the seat of new tumors. 

The blood vessels sometimes transport the fungus, 
as has been established in man, in consequence of the 
ulceration of the jugular vein in contact with a dis- 
eased focus. 

Generalization does not appear to take place in ani- 
mals, perhaps on account of the premature slaughter 
of the subjects, a course which is most frequently 
followed and which is the most economical. Acti- 
nomycosis has, however, been encountered in the 
spleen in cattle and in the muscles in pigs.* 

Besides the functional disturbances to which they 
give rise (disturbances of mastication, deglutition, 
respiration, etc.) the actinomycomata, especially when 
they have ulcerated, cause a progressive emaciation 
of the diseased animals. 

Botryomycosis. 

In certain indurations of the testicular cord of the 
castrated horse a special parasite is found to which 
the various names of botryomyces (Bollinger), dis- 
comyces (Rivolta), botryococcus ascoformans (Kitt), etc., 
have been given. It has also been met with in cer- 
tain forms of fistulous withers, in tumors at the point 
of the shoulder, and in certain nodosities of the skin 
and subcutaneous tissue, etc. Czoker has also re- 

* [The actinomyces-like parasite found in the muscles of swine 
(Actinomyces musculorum suis) is not identical with the Acti- 
nornyces bovis sive hominis. C. Giinther; Bakteriologie, p. 
327. P.] 




Microbic Diseases Individually Considered. 281 

corded its occurrence in the pus of a chronic mam- 
mitis in the cow. 

The parasite appears in the form of a cluster of 
small spheres full of micrococci. Fi 

The latter are united by a gelatinous 
substance, and each of the spheres 
is inclosed by a double contoured 
membrane. New spheres develop 
at the periphery of the colony. The 
parasite is related to the genus as- 
COCOCCUS. Botryococcus asco- 

Staining may be obtained by the forma f Sli s htl y 

J " . J magnified. Kitt. 

method of Gram, with eosm as con- 
trast stain. The eosin becomes fixed especially on 
the gelatinous fundamental substance ; picric acid 
has a similar action. 

The pathogenesis of the lesions (botryomycomata, 
mycofibromata, mycodesmoides) is the same as that 
of the actinomycomata. They are inflammatory 
tumors of slow evolution showing a mixture of 
granulation tissue and fibrous tissue of different de- 
grees of condensation. The granulation tissue is 
often arranged in tuberculiform masses of which the 
parasite occupies the center, and the elements of 
which may suppurate, become necrosed, and give rise 
to fistulous tracks. 

The parasite generally confines its ravages to its 
primary seat ; it may, however, migrate toward the 
lymphatic glands and may even pass into the blood. 
Metastatic foci then appear, commonly localized in 
the lung, skin, etc. In the lung these lesions have a 
resemblance to those of glanders. 
24 



282 Manual of Veterinary Microbiology. 

The germ gains entrance to the system through 
operative (castration) or accidental wounds. 

Bovine farcy. 

This disease is peculiar to the bovine species. 
Formerly noticed in France, it appears to be much 
rarer at the present time ; it exists in the Guadalope. 
It is usually located on the limbs, manifesting itself 
by painless cords along the course of the subcuta- 
neous veins and terminating at the corresponding 
lymphatic glands; the brachial, prescapular and pre- 
pectoral glands are those most frequently attacked. 
These cords and glands may suppurate; the suppura- 
tion is always slowly evolved and the whole assumes 
the characters of a cold abscess, with very thick and 
indurated walls. After these abscesses are opened 
the subject seems to recover, but very soon other tu- 
mors appear. The animal pines away and dies by slow 
decline. The autopsy brings to light pseudo-tubercu- 
lous lesions with purulent centers in the internal 
organs : lungs, liver, spleen, and lymphatic glands. 

We are indebted solely to M. Nocard for the bacte- 
rial study of the disease. 

Microbe. There is found in the pus of the abscesses 
and in the center of the pseudo-tubercular lesions " a 
long, slender bacillus, appearing under 
the form of small, intricately interwoven 
masses, the central part forming an 
opaque nucleus from which radiate to 
Bovine farcy, the periphery myriads of fine prolonga- 
(M. and L.) tions, the majority of which seem to 
branch." The dimensions of this bacillus are nearly 
the same as those of the bacillus of rouget. 




Microbic Diseases Individually Considered. 283 

The microbe of cattle farcy is purely aerobic. 

Cultures. Cultures are easily obtained between 30 
and 40. In bouillon it forms rounded pellicles of a 
dull gray color and oily appearance, floating in the 
liquid. On agar and gelatin it forms small masses 
more or less regularly rounded, opaque, and thicker 
at the periphery than at the center. On potato the 
growth is rapid and takes the form of dry salient 
plates, often depressed at their center. 

The microbe reproduces itself in cultures in the 
form of star-like masses of intertwining filaments, 
and it is only at the periphery that the bacillar nature 
of its constituent elements can be recognized. Ac- 
cording to M. Nocard, the ramifications observed in 
these masses are false ; the bacillus divides trans- 
versely, then the terminal filament inclines to a right 
angle, allowing its generator to continue its direct 
course. The organism, therefore, is not a true clado- 
thrix. 

Research and coloration. The bacillus of cattle farcy 
is easily colored; it is decolorized by the reaction of 
Gram when the contact with the alcohol is too pro- 
longed, but it takes the double stain of Weigert per- 
fectly. The spores are stained with difficulty. 

Experimental inoculations. The disease is inocula- 
ble to cattle, to the sheep and to the guinea pig; not 
to the rabbit, cat, dog, nor horse. 

As a result of intra-peritoneal inoculation in the 
guinea pig the serous lining becomes studded with 
tuberculiform nodules, in the purulent center of 
which the characteristic tufted bacilli are contained ; 
these nodules are especially abundant in the omen- 
turn, which itself has become much enlarged. The 



284 Manual of Veterinary Microbiology. 

abdominal viscera are altered only in their serous 
covering, the tissue proper being unaffected, whilst 
the thoracic organs are exempt from lesions. 

The injection of virulent material into the veins is 
followed by a generalized pseudo-tuberculosis, the 
bacilli being found in tufts in the center of all of the 
lesions. 

In both cases death occurs from the ninth to the 
twentieth day. Subcutaneous inoculation in the 
guinea pig always produces a voluminous local ab- 
scess ; the corresponding lymphatic glands also sup- 
purate and recovery takes place only after a period 
of great emaciation, leaving behind it an induration 
of the lymphatics. Generalization may, however, be 
occasionally observed. 

Intra-vascular inoculation in the sheep and in cat- 
tle also produces nodular lesions distributed through- 
out the viscera, but death does not immediately fol- 
low, the disease developing Very slowly. 

Hypodermic inoculation in the same animals pro- 
duces an abscess which ulcerates on different occa- 
sions, seems to heal, then reappears later, exactly as 
in the natural disease. 

Subcutaneous inoculation in refractory animals oc- 
casions an abscess which quickly heals, 

Tetanus. 

Tetanus occurs spontaneously in all the domesti- 
cated animals and in man ; it is more frequent in the 
horse, ox, sheep, and goat, but it has also been ob- 
served in the pig and dog. 

The contagiousnesss of tetanus, which had long 
been suspected from clinical observation, has been 



Microbic Diseases Individually Considered. 285 

demonstrated by experimental inoculation, and its 
causative agent has been brought to light by bacteri- 
ological research. 

Microbe. The bacillus of tetanus, first described 
by Nicolaier, is a slender rod, measuring 3/* to 5^ in 
length, homogeneous, or containing at one end an 
enlargement in which appears a spherical spore of a 
diameter double .or quadruple that of the filament. It 
has, therefore, when sporulated, the form of a pin, 
of which the brilliant, lustrous spore represents the 
head. In wounds the bacillus sometimes attains a 
length much greater than that indicated above. 

The bacillus of Nicolaier is motile, -pig. 17. 

its movements resembling those of the ^ t / 
septic vibrio, but these movements * *& 
cease when fructification is accom- f ^'j t 
plished. It is anaerobic, multiplying / * 

only when the atmospheric air is ex- Bacillus of te- 
eluded. A rarefied atmosphere- is, ^JTot 
however, compatible with its vitality, " sporulated. (M. 
and this will account for its mutlipi- and L.) 
cation in free media, the aerobic germs protecting it 
from contact with too large a proportion of oxygen. 

The tetanus bacillus remains intrenched in the in- 
fected focus and does not penetrate into the blood 
during life; at the approach of death, or a certain 
time after death, it may be found in remote parts in 
which the deoxygenation of the blood has allowed of 
its multiplication. 

The bacillus is met with in its two forms, homo- 
geneous and sporulated, in the pus of wounds which 
have given rise to tetanus, the sporulated form, how- 
ever, being less abundant than the non-sporulated. 



286 Manual of Veterinary Microbiology. 

In young cultures the latter form also predomi- 
nates. 

Action of physical and chemical agents. The spores 
are very resistant to heat ; they support a tempera- 
ture of 80 during six hours, or 90 during one hour. 
In moist heat at 100 they are killed in a quarter of 
an hour, at 115 in five minutes. Spores which are 
thoroughly dried retain their vitality for a long time 
if protected from the light. Exposed to the light 
they do not survive more than one month especially 
if, at the same time, exposed to the air. 

The bacilli of Nicolaier are little sensitive to the 
action of antiseptics, notably less so than the septic 
vibrio. Thus, sublimate solution, at 1 to 1000, only 
kills these bacilli after three hours; three per cent 
carbolic acid solution after ten hours. The spores re- 
sist a five per cent solution of carbolic acid for fifteen 
hours, the same solution with an addition of 0-5 per 
cent of hydrochloric acid, two hours, and 1 to 1000 
sublimate solution with a like proportion of hydro- 
chloric acid, thirty minutes. They are unaffected by 
the gastric juice and by putrefaction; active cultures 
having been administered to rabbits, guinea pigs, mice 
and dogs by way of the mouth, all these animals re- 
mained healthy, whilst their excrements were virulent 
for other animals. 

Cultures. The bacillus of Nicolaier vegetates in 
artificial culture media when the oxygen of the air is 
excluded and when under suitable conditions of tem- 
perature; culture vessels should therefore be em- 
ployed in which a vacuum has been created or in 
which the atmosphere has been replaced by hydro- 
gen, carbonic acid, etc. Solid liquetiable media, such 



Microbic Diseases individually Considered. 28? 

as gelatin and agar, are sometimes freed from air by 
boiling for half an hour; the mass is solidified by 
rapid cooling, inoculated by deep puncture, and the 
surface of the medium covered with a layer of ster- 
ilized oil which prevents the absorption of the at- 
mospheric oxygen. Deoxygenation can also be ac- 
complished by the addition to the same media of sub- 
stances which readily absorb oxygen. Kitasato ad- 
vises the addition to gelatin or agar, of glucose two 
per cent, sulpho-indigotate of soda 0*1 per cent, blue 
turnsol live per cent. 

The most favorable temperature varies between 
38 and 39, but multiplication still occurs at 18 and 
continues even at 43. 

The natural inoculation substances at our disposal : 
pus of wounds which have given rise to tetanus, earth, 
forage, etc., are always contaminated with other germs 
than those of this disease ; hence, impure cultures 
are first obtained from which the bacillus of Nicolaier 
must be isolated. With this aim we take advantage 
of the great resistance of its spores, and heat the 
sporulated cultures in a water bath at 80 to 90 for 
three quarters of an hour to one hour ; traces of 
these cultures are sowed in anaerobic tubes which are 
then closed over the flame and the gelatin spread out 
on their walls by a rotatory motion. 

The colonies which then develop are composed ex- 
clusively of tetanus bacilli, as may be proved by. in- 
oculation. If some spores of the septic vibrio have 
resisted the heat and produced a collateral vegetation, 
we should then resort to the action of antiseptics ; 
1 to 1000 sublimate solution kills- the tetanus bacillus 



288 Manual of Veterinary Microbiology. 

only after three hours, 3 per cent carbolic solution 
only after two hours. (Sanchez-Toledo.) 

In bouillon the development is very rapid ; the 
liquid becomes turbid in one day and sets free small 
bubbles of gas ; the growth abates and becomes de- 
posited toward the fifteenth day. 

When a gelatin tube is inoculated by deep punc- 
ture and kept at 18, the growth forms, after four or 
five days, small cloud-like points from which fine lines 
radiate perpendicular to the puncture. The culture 
has a floculent appearance, slowly fluidifies the gela- 
tin, and bubbles of gas are disengaged; when the 
gelatin is entirely fluidified the culture is deposited 
in the form of white flakes. 

Cultures upon agar are less characteristic. Growth 
also takes place Upon serum and potato. Cultures 
of tetanus emit a smell of burnt horn and produce 
various gases, including carbonic acid and hydro- 
carbons. 

Research and coloration. The bacillus of tetanus is 
easily stained with the aniline colors ; it takes the 
Gram very well and appears as a slender rod, uniform 
in size or swollen at one of its extremities. Before 
the formation of the spore this enlargement stains 
like the filament itself. The spores are not stained 
by this process but may be colored by the method 
usually employed for the staining of spores. 

Experimental inoculations. The disease is inocula- 
ble to the smaller animals and notably, with a sus- 
ceptibility decreasing in the order in which they are 
named, to the mouse, white rat, guinea pig, rabbit, dog, 
pigeon and chicken. The mouse, white rat and guinea 
pig are extremely susceptible, 0.002 cub. cent, being 



Microbic Diseases Individually Considered. 289 

sufficient to produce in these animals the typical dis- 
ease with fatal results in 36 to 40 hours. The rabhit 
requires 10 to 30 drops ; the symptoms appear from 
the second to the third day and death occurs four to 
ten days later. The dog, pigeon and chicken are 
less susceptible and sometimes survive large doses. 

The inoculation succeeds well in the connective 
tissue, in the peritoneum and in the arachnoid. In 
the connective tissue it causes an oedematous swell- 
ing, the contractions appearing first of all in the ad- 
jacent muscles. When inoculated in the peritoneum 
or in the blood these appear siiniltaneously in all 
parts of the body. The disease is not produced either 
by inhalation or ingestion of virulent products. 
Ingestion of tetanic toxines is also without effect, 
these toxines being destroyed by the digestive juices. 

According to the dose inoculated and the suscepti- 
bility of the animals the experimental disease may be 
acute or chronic, fatal or curable. 

In connection with experimental inoculations three 
cases must be distinguished. The material to be in- 
jected may consist of the entire culture, of the amor- 
phous part alone freed from its microbes by filtration, 
or, finally, of the microbes alone deprived of their 
soluble products by filtration, lixiviation, or heat. 

In the first two cases inoculation of the entire 
culture or of its soluble products the classic disease 
is reproduced. In the last case, according to MM.Vail- 
lard and Vincent, the inoculation remains without ef- 
fect except when excessive doses are employed. The 
tetanus bacillus, therefore, appears to be unable to 
multiply in the organism in the absence of its toxines, 
25 



290 Manual of Veterinary Microbiology. 

being destroyed, in such case, by the phagocytes; it 
excites an active diapedesis which may be observed 
after injections into the anterior chamber of the eye. 
The toxines of cultures, on the other hand, seem to 
repel the leucocytes and thus protect the microbes 
from their destructive action. Certain other sub- 
stances may take the place of this action : inoculation 
of spores alone is followed by tetanus when lactic 
acid, diluted to 1 to 500,'or trimethylamin, is injected 
at the same time, or when a contusion is produced in 
the inoculated tissues. Simultaneous injection of the 
germs of tetanus a.nd the micro-bacillus prodigiosus 
also allows the irruption of the disease; the prodigio- 
sus attracts the phagocytes to itself and thus forms a 
barrier behind which the tetanus bacillus multiplies 
and secretes its protective toxines. 

The experiments of Yaillard and Vincent have been 
repeated by Sanchez-Toledo, who seems to have suc- 
ceeded in transmitting the disease by means of cult- 
ures freed from their toxines and from all adjuvant 
substances. 

Etiology and pathogeny. The tetanus bacilli or their 
spores exist in the soil; the disease has often been 
produced by inoculation of water with which certain 
specimens of soil have been washed. From the soil 
they are transported in the dried forage, pass un- 
changed through the digestive canal of herbivora and 
with the manure are returned to the soil. By inocu- 
lations they have been demonstrated in hay and in 
the excrements of healthy horses and cattle. Tetanus 
is therefore of telluric origin ; but it can also be trans- 
mitted from one animal to another, either directly or 
indirectly. The microbe being inactive when inhaled 



Microbic Diseases Individually Considered. 291 

or ingested, it must gain entrance to the system 
through a solution of continuity. 

All wounds are not alike adapted to the growth of 
the tetanus bacillus ; the germ is anaerobic and conse- 
quently requires a medium little accessible to atmos- 
pheric air ; besides, it is inoffensive if not protected 
against the phagocytes by accessory conditions. The 
production of tetanus, therefore, as a rule, requires 
deep, anfractuous, contused wounds or those contami- 
nated by other germs, and more especially by the 
ordinary pyogenic species. Nevertheless, tetanus has 
often been observed to follow insignificant wounds. 
It should be observed that wounds favorable for the 
growth of the bacillus of tetanus are also favorable 
for that of the septic vibrio. Now, the bacillus of 
malignant oedema is also found in tetanogenic earth, 
so that the same wound may be contaminated with 
both germs ; gangrene, however, running its course 
more rapidly than tetanus, the latter may only appear 
after recovery from the former, or may not appear 
at all if the subject succumbs to the septicaemia 
(Verneuil). 

The bacillus of tetanus secretes in the wound spe- 
cial toxines which have a poisonous action on the 
organism similar to that of strychnine. If a culture 
of tetanus be filtered so as to completely separate the 
microbes from the soluble part, and the latter be in- 
jected to animals, an absolutely typical tetanus re- 
sults. This filtered liquid is very toxic : one-twentieth 
of a drop kills a mouse in thirty-six hours, one drop 
kills a guinea pig in twenty-four hours. 

The nature of the tetanic poison is yet incom- 
pletely known in spite of numerous investigations. 



292 Manual of Veterinary Microbiology. 

Brieger isolated from impure cultures on meat several 
toxic ptomaines : tetanin, tetanotoxin and spasmatoxin. 
The first gives the typical tetanus when injected in 
very small doses, the second produces tonic and clonic 
convulsions, and the third hyper-salivation and con- 
vulsions. According to more recent investigations 
the tetanic poison appears to be an albumin related 
to the diastases or soluble ferments. It is destroyed 
by a temperature of 65 in five minutes, is insoluble in 
alcohol, soluble in water, and adheres to precipitates 
of alumina and phosphates. It is destroyed by heat 
and preserved by desiccation if this is rapidly ob- 
tained in a vacuum and without the aid of heat. 

This toxine appears to act more particularly on the 
muscular tissue, which would explain the appearance 
of the first tetanic contractions in the muscles which 
are adjacent to the wound or to the place of inocula- 
tion. When administered by way of the digestive 
canal it is inactive. 

From the preceding considerations it may be in- 
. ferred that the microbe of Nicolaier produces its ef- 
fects only by means of the diastases which it secretes. 
Moreover, it gains entrance into the blood only in 
the last moments of life, or after death. 

Attenuation. Vaccinations. By heating the filtrate 
to different temperatures between 55 and 100 until 
it becomes inactive, and inoculating this material to 
mice, we determine a mild, curable form of tetanus, 
but one which is not followed by immunity. 

Kitasato endeavored to obtain immunity against 
tetanus by the action of trichloride of iodine. He 
injected 0-3 cc. of filtered culture under the skin of a 
rabbit and immediately afterward, in the same place, 



Microbic Diseases Individually Considered. 293 

3 cc. of 1 per cent solution of trichloride of iodine, 
the latter injection being repeated every twenty-four 
hours until the rabbit had received in all 0-15 grams 
of the trichloride. If, after fourteen, eighteen, twenty- 
five days, 2 cc. of the virulent filtrate, or 2 to 3 cc. of 
virulent bouillon culture, be injected, the tetanic 
symptoms which supervene disappear in a few days 
and the animal can then receive an injection of 5 cc. 
of virulent culture without manifesting any symp- 
toms. The rabbit is therefore vaccinated; but this 
process is inconstant, and sixty per ceiit of the rabbits 
do not obtain immunity. The mouse and guinea pig 
do not obtain it at all. Now, the serum and blood 
of these vaccinated rabbits possess the very interest- 
ing property of destroying the toxine; they are toxin- 
icidal. A mixture, twenty-four hours old, of 1 cc. of 
a virulent culture with 5 cc. of this serum can be in- 
jected with impunity to mice. It is even possible 
by means of this serum to check the disease experi- 
mentally developed in the mouse. Further, the in- 
jection of O2 .to 0*5 cc. of this serum into the peri- 
toneum of the mouse confers an immunity of from 
forty to fifty days' duration. 

Tizzonr and Cattani have obtained immunity in the 
dog and pigeon by the injection of progressive doses 
of cultures of gradually increasing virulence. The 
serum of the dog, thus vaccinated, is toxinicidal and 
confers immunity on the dog and on the mouse but 
not on the rabbit or guinea pig, and does not check 
the disease when already established. These authors 
have isolated and obtained in the dry state the tox- 
inicidal product of the serum (antitoxin) ; they appear 



294 Manual of Veterinary Microbiology. 

to have successfully used it in the treatment of human 
tetanus. 

Vaillard, by injecting into the blood of rabbits, 
first, in several doses, 40 cc. of filtered culture heated 
to 58, second, 10 cc. of filtered culture heated to 51, 
and, finally, 15 cc. of filtered culture not heated, did 
not immunize these animals, but communicated to 
their blood the toxinicidal power; hence it follows 
that there is no relation between the toxinicidal 
property and the refractory state.* 

* [Of the more recent investigations bearing on the etiology of 
tetanus and the production of immunity, the following points 
may be briefly noted : 

The necessary co-operation of other microbes at the place of in- 
fection (in the naturally acquired disease) has been reaffirmed. 
(Vaillard and Rouget.) 

According to Courmont and Doyon, the special tetanus toxine 
results from a fermentation excited in certain tissues of the or- 
ganism by a soluble ferment secreted by the bacillus tetani. 
(Compt. rend. Soc. de Biologie, March, 1893.) 

Immunity can be obtained by repeated injection of cultures of 
the bacillus tetani grown in bouillon prepared from the thymus 
gland of calves, or of filtered cultures to which a certain propor- 
tion of an extract of this gland has been added (Brieger, Kitasato, 
etc.) ; the blood serum of animals thus immunized possesses im- 
munizing properties. By repeated injection of gradually increas- 
ing doses of virulent cultures to animals (rabbits, dogs, horses) 
which have been thus immunize^, the protective power of the 
serum of these animals becomes greatly increased. This serum 
then furnishes an effective " vaccine." The serum of such im- 
munized animals or a preparation from the same (anti-toxin : Tiz- 
zoni, Cattani) has been used (by repeated subcutaneous injection 
of considerable doses) in the treatment of tetanus in man, and 
apparently with good results. (Ref . Centralbl /. Bacteriologie, XIII, 
4,14; XIV, 4, 12,19; .XT, 4.) 

The curative action of the serum of immunized animals de- 
pends upon its immunizing properties; it localizes the tetanus 



Microbic Diseases Individually Considered. 295 

Diphtheria. 

In its widest meaning this term is applied to a 
special form of inflammation of the integuments in 
which a concrete exudate is produced in the thick- 
ness of the derm and involves its mortification ; this 
exudate is called diphtheritic. When the fibrinous 
deposit is limited to the epithelium the inflammation 
is said to he croupous. These two forms of inflamma- 
tion can be produced by very varied causes : these 
are mechanical (compression), or physical (burns), or 
chemical (caustics), or, finally, biological (parasites 
and microbes). Among the number of parasites may 
be mentioned the gregarinse of the contagious epi- 
thelioma of poultry and the coccidia of typhlitis of 
the same animals; among microbes a large number 
possess the same faculty ; of these we need only re- 
call the diphtheritic exudates of pneumo-enteritis of 
the pig, of acute glanders, and of petechial typhus, 
etc. Diphtheria of wounds, or hospital gangrene, 
characterized by a superficial necrosis of the divided 
tissues, must also be attributed to the contamination 
of the latter by micro-organisms. 

From a clinical point of view the term diphtheria re- 
fers to a specific disease due to a special germ, and 
manifesting itself by an inflammation, with croupous 
or diphtheritic evolution, of the respiratory and some- 

by protecting the parts of the nervous system not yet attacked by 
the tetanus poison. (Tizzoni, Cattani: Id. XV, 17.) 

Tests of the curative action of the serum of immunized animals 
made by Nocord, in the case of two sheep artificially infected 
with tetanus, resulted unfavorably. (Ref. Jour. Comp. Path. VII, 



296 Manual of Veterinary Microbiology. 

times digestive mucous membrane. This disease 
has been thoroughly studied in man ; it also occurs 
in certain species of animals, particularly in birds and 
in the calf. At one time it was believed that human 
arid aviau diphtheria were identical, but this view is 
no longer entertained. The 'two diseases have a 
different evolution and different microbes. 

Human diphtheria. Klebs and Loffler discovered in 
the false membranes a bacillus, straight or curved, 
with rounded ends, sometimes club-shaped, at other 
times less than the average thickness. It meas- 
ures 2*5// to 3/z in length by 0*7// in thickness; it is 
especially aerobic but also grows when the air is ex- 
cluded; it is met with in the superficial zone of the 
false membranes. The germ grows on most of our 
artificial media; it is stained by Loftier' s methylene 
blue and by the Gram method. Roux and Yersin 
have completed its biological study; they isolated it 
by inoculating a series of serum tubes without re- 
charging the platinum wire : the colonies of diph- 
theritic bacilli appear as grayish-white rounded 
growths with the center more opaque than the 
periphery. 

Excoriated mucous membranes, inoculated with a 
culture, soon show the characteristic false membrane 
in rabbits, guinea pigs, cats, pigeons, and chickens. 

The subcutaneous injection of a few drops of the 
liquid kills the guinea pig in thirty-six hours with 
general vascular dilatation and pleural effusion. The 
rabbit requires 1 cc. of the culture and, like the pigeon, 
dies after several days. The dog and the sheep also 
succumb. In all cases a hemorrhagic oedema is pro- 
duced at the place of inoculation and the vessels of 



Microbic Diseases Individually Considered. 297 

the different organs are congested. When' death of 
the inoculated animals is delayed they exhibit paraly- 
sis resembling that observed in the child suffering 
from this disease. 

The bacilli occur only in the specific lesions ; there 
they secret a poison the absorption of which deter- 
mines the general symptoms of diphtheria. Cultures 
freed from microbes by filtration are very toxic ; un- 
der their influence guinea pigs exhibit a pronounced 
dyspnoea; rabbits are attacked with progressive 
paralysis and often with diarrhoea. The dog and the 
sheep also succumb after showing symptons of paral- 
ysis. 

The toxic substance of cultures is a diastase pre- 
senting much analogy with that of tetanus ; in the 
digestive canal it is innocuous. 

Behring appears to have obtained immunity in 
guinea pigs : 1st, by inoculating them with cultures 
sterilized at 65 to 70 C.; 2d, by injecting them with 
a mixture of one part of trichloride of iodine and 
five hundred parts of these same cultures ; 3d, by in- 
oculating them, with the serous or sanguinolent 
liquid taken from the pleura of guinea pigs dead of 
diphtheria. 

Avian diphtheria. Birds are very subject to a dis- 
ease manifesting itself by the production of false 
membranes and diphtheritic exudates on the mucous 
membranes of the mouth, pharynx, oesophagus, nose, 
eyes, larynx, trachea, lungs, air cavities, intestines, 
and upon the skin. This disease is very contagious 
although much less severe than that of man ; like the 
latter it may occasion rapid mechanical asphyxia, but 
more frequently it is protracted, presenting remis- 



298 Manual of Veterinary Microbiology. 

sions and exacerbations, and leading to the emaciation 
of the affected birds. A catarrhal inflammation 
which sometimes occurs in the attacked mucous 
membranes, by provoking glairy morbid secretions, 
aggravates the cachexia and hastens the fatal termina- 
tion. 

Loffler attributes the disease to a special bacillus, 
staining by the methods of Gram and Weigert, which 
he met with in the fluid products of the inflamed 
mucous membranes, in the false membranes, the 
lesions of the liver, and in the blood. This bacillus 
has nearly the same dimensions as that of human 
diphtheria but it is smoother and more uniform in 
thickness. It is abundantly present in the superficial 
layer of the false membranes, rare or absent in the 
deep layers. Along with this specific bacillus other 
microbes are constantly found in the concrete exu- 
dates. 

Loffler reproduced avian diphtheria in the rabbit 
and the pigeon; death, when it occurred, supervened 
less quickly than with the human bacillus. The 
guinea pig and the dog are also more sensitive to the 
latter bacillus than to that of birds. Loffler culti- 
vated and described the microbe and with it repro- 
duced the disease in birds and the rabbit by inoculation 
of cultures on the mucous membrane, on the skin, 
and in the subcutaneous cellular tissue. 

The disease is propagated by means of the morbid 
matters (discharge, faeces, false membranes) of the dis- 
eased animals. The virus gains entrance into the 
organism of healthy animals with the food or by the 
air. Contamination most frequently takes places in 
an indirect manner but may also occur directly. M. 



Microbic Diseases Individually Considered. 299 

Meguiu has recorded an outbreak in pigeons in which 
the young animals quickly succumbed to diphtheria; 
he attributes the disease of the young pigeons to the 
fact that the mothers were affected with old, little 
marked oasophageal lesions the virulent products 'of 
which mingled with the lactescent liquid of the crop 
were fed directly to the young. 

The disease appears in poultry pens in consequence 
of the importation of birds coming from infected cen- 
ters or on the return of animals which have been ex- 
hibited in bird shows. The virus appears to be 
preserved for a considerable time in contaminated 
pens and in manure. 

In spite of the cases advanced by different observ- 
ers it appears to be well established that the disease 
of fowls is not transmissible to man. 

Diphtheria of calves. There exists in the bovine 
species a diphtheritic affection transmissible chiefly to 
calves and also having its origin in a special bacillus. 
It localizes itself in the back of the throat, in the 
trachea, and in the bronchi. 

Rabies. 

Rabies occurs, under natural conditions, in all do- 
mesticated animals and in several wild species : wolf, 
fox, jackal, bear, etc. It is very much the most fre- 
quent in carnivora, and more especially in the dog 
and wolf. 

Microbe. In despite of numerous researches con- 
cerning the pathogenic agent of rabies, its morpho- 
logical characters are yet unknown. Its existence 
however can not be doubted; it multiplies in the 
organism to which it is inoculated, and loses or gains 



300 Manual of Veterinary Microbiology. 

in virulence like the well defined microbes of other 
diseases. Fol and Babes have each described micro- 
cocci which they were able to cultivate in bouillon, 
and with which cultures they believed that they trans- 
mitted rabies. Babes also describes a short bacillus. 
Mottet and ProtopopofF isolated from the brain very 
fine bacteria, cultures of which in bouillon, according 
to these authors, gave the typical disease. 

Action of physical and chemical agents. The viru- 
lence of an emulsion of the spinal cord of a rabid 
animal is dissipated when heated twenty-four hours 
at 45, one hour at 50, and half an hour in steam at 
100. 

It resists a temperature of 20 for thirty hours at 
least. It is destroyed by fourteen hours exposure at 
37 to solar light, Desiccation in the air rapidly di- 
minishes its virulence and destroys it in a few days. 

Virulence is retained in cadavers for several weeks 
if decomposition is prevented, but it is destroyed by 
putrefaction. 'Under the influence of 1 to 1000 sub- 
limate solution, 2 or 5 per cent solution of perman- 
ganate of potash, or 50 per cent alcohol, the virus is 
quickly impaired; 15 percent alcohol, on the other 
hand, preserves it intact for seven days at least. 
Even a large dose of the emulsion proves inoffensive 
when it has been rendered either acid or alkaline. 
Perfectly neutral glycerin (30 B). preserves its patho- 
genic power in a perfect manner. 

Experimental inoculations. With the exception of 
some individuals which are naturally insusceptible, 
the majority of the mammalia contract rabies when 
inoculated. 

The species used in the laboratories are the rab- 



Microbic Diseases Individually Considered. 301 

bit, guinea pig, dog, ape, rat, and birds. The last 
named species almost invariably recover spontane- 
ously (Gibier). The virulent material is taken from 
the nervous substance (brain, medulla, cord and 
nerves), from the salivary glands, the saliva, bron- 
chial mucus, and from the pancreas. The milk occa- 
sionally shows itself virulent, but the blood is not; 
complete transfusion of blood from a rabid dog to a 
healthy subject did not produce the disease (Bert.) 
In practice we resort more especially to the nerve 
centers, which furnish a pure virus; for use, a parti- 
cle is reduced to pulp and the latter diluted in steril- 
ized water or bouillon ; in this way a white milky 
fluid is obtained, a veritable emulsion of the nervous 
substance. 

The inoculation maybe performed in various ways; 
it may be subcutaneous, intra-muscular, intra-venous, 
intra-ocular, intra- cranial, intra-nervous, etc., the re- 
sult varying according to the method employed. 

In the dog, subcutaneous and intra-venous inocula- 
tions are usually followed by dumb rabies, without 
barking or fury. Furious rabies can be obtained by 
the same methods, but with small doses of virus. 
The smaller the amount of the virus employed the 
more readily is the furious form of the disease ob- 
tained, the period of incubation being at the same time 
prolonged. The injection of very small doses maybe 
ineffective in the dog without conferring immunity, 
whilst inoculation of large quantities of virus may 
give immunity without the manifestation of any 
symptoms of rabies. 

Intra-muscular inoculation is followed by rabies 
with more certaintv than subcutaneous inoculation. 



302 Manual of Veterinary Microbiology. 

In the dog, intra-cranial inoculation is always fol- 
lowed by furious rabies; the-period of incubation for 
the virus of the streets is from fourteen to fifteen 
days, on an average. 

The inoculation may also be made in the nerve 
trunks, a previous lesion of the nerve fibers increas- 
ing the chances of success; in this method the period 
of incubation is not the same in all cases, and is 
always logger than by trephining; the virus vege- 
tates in the nerve and progresses from the periphery 
toward the center, the disease being more tardy in 
its appearance as the course to be traversed by the 
virus is more extended. Inoculation in the posterior 
nerve trunks is followed by paralytic rabies; in the 
anterior trunks by furious rabies. 

The insertion of the virus into the anterior cham- 
ber of the eye is invariably followed by rabies, the 
incubation in this case being thirteen to sixteen days 
for the natural virus of dogs. 

In the rabbit, intra-cranial inoculation enables us to 
make some important observations : the incubation 
is from fifteen to seventeen days, and death occurs in 
the course of the next four days. The disease mani- 
fests itself by paralytic phenomena with progressive 
course ; rabies of the rabbit is, therefore, dumb rabies. 
However, cases of furious rabies are also -observed, 
and Ferre has shown that symptoms of excitement 
(accelerated respiration, etc.) precede the paralytic 
symptoms. 

Passage- of the virus of natural rabies from the first 
rabbit to a second, from the second to a third, and so 
on, exalts the activity of the rabic virus. This exal- 
tation shows itself in the shortening of the period of 



MicroUc Diseases Individually Considered. 303 

incubation which becomes abbreviated to eight days 
after twenty-five passages and to seven days after 
fifty passages. The virus is then 'acclimated in the 
rabbit, which dies regularly after seven days what- 
ever generation the virus employed may belong to ; 
the virus of rabies thus exalted takes the name of 
fixed virus. It is exalted not only for the rabbit but 
also for the dog itself, which always contracts the dis- 
ease from intra-vascular inoculation of this virus, 
whilst similar inoculation of natural virus is uncer- 
tain in its results. 

In the guinea pig, which readily takes rabies by 
intra-cranial inoculation of natural virus, virulence is 
exalted by passage in series, until, after the eighth 
inoculation, it becomes fixed. The incubation period 
is then five days. Rabies in the guinea pig often 
shows a distinct period of excitement; the fixed virus 
of the guinea pig is more active for the dog than the 
natural virus. 

In the ape, passage in series attenuates the viru- 
lence ; whilst the virus taken from the first ape kills 
the rabbit in thirteen to sixteen days, that of the sec- 
ond allows an incubation in the rabbit of fourteen to 
twenty days, and that of the sixth, thirty days. In- 
tra- venous inoculation of the virus of the sixth gen- 
eration no more produces the disease in the dog, and 
even intra-cranial inoculation is uncertain. The viru- 
lence attenuated in the ape can be restored to its 
original activity by a series of passages through 
rabbits. 

Etiology and pathogeny.R&b\ea is undoubtedly 
caused by a living virus. It is transmitted from one 
animal to another by direct contagion. Mediate 



304 Manual of Veterinary Microbiology. 

transmission seems to be of very rare occurrence. 
The virus is preserved only for a very short time in 
external media, a circumstance which diminishes the 
chances of its transportation; M. Galtier, however, 
has pointed out the possibility of infection by the in- 
tact ocular mucosa, and it is quite conceivable that 
this might actually occur in man through the projec- 
tion or transference of a virulent liquid to the eye. 
Contamination by ingestion of the flesh coming from 
a diseased animal appears to have occurred a certain 
number of times in the dog (Zundell). Experimental 
tes.ts of this mode of infection have, however, given 
only negative results. 

The disease is transmitted to the healthy individual 
in nearly all cases by the bite of a rabid animal, such 
bite being equivalent to an inoculation with virulent 
saliva. Bites of carnivora (dog, cat, wolf) are the most 
dangerous, on account of the severity of the wounds 
which they inflict ; those of the horse and deer are 
less dangerous; bovines have not as yet been known 
to communicate the disease. MM. Nocard and Roux 
have shown that the virulence appears in the saliva, 
on an average, twenty-four hours before the first 
symptoms, and that there are cases in which it ap- 
pears one or two days earlier. On this account they 
recommend that all dogs which have bitten other 
animals or human beings should be kept under ob- 
servation for three days at least before certifying to 
their condition. 

Bites which involve the nerves or muscles are 
among those which are the most likely to communi- 
cate rabies. 

Besides bites, direct inoculation may occur in the 



Microbic Diseases Individually Considered. 305 

course of an autopsy. Inter-placentary transmission 
has not been established ; cases of such transmission 
which have been brought forward have not been 
verified. 

The period of incubation is very variable ; the dis- 
ease most frequently appears in the course of the 
first two months after the bite, but it may appear 
after a few days as well as after several months. 
These differences depend upon the more or less easy 
propagation of the virus. Absorption takes place 
chiefly by the nerves ; in fact, inoculation in a nerve 
trunk induces rabies more rapidly than its insertion 
in the connective tissue at a corresponding point, al- 
though less quickly than when introduced into the 
cranial cavity. The virus, therefore, seems to vege- 
tate in the nerve and progress toward the cerebro- 
spinal center, thence radiating along the nerves. 

After inoculation in the sciatic the primary rabic 
symptoms indicate changes in the lumbar cord, the 
cervical cord and the meclula being only attacked at 
a later stage. In one case in the guinea pig the dis- 
ease was limited to paraplegic symptoms alone; by 
interrupting the continuity of the spinal cord at the 
level of the dorso-lumbar region the dorso-cervical 
cord was protected from the action of the virus. 
Some cases in man which have been carefully ob- 
served support to some extent this view as to the 
propagation of the virus of rabies: after a bite on the 
right arm the nerves of this arm, inoculated along 
with those of the left arm, alone showed themselves 
virulent. Incubation should therefore be shorter in 
proportion as the length of nerve to be traversed be- 
26 



306 Manual of Veterinary Microbiology. 

fore arriving at the ceriter is itself shorter. How- 
ever, under natural conditions the duration of this 
latent period is influenced by various factors, such as 
the quantity of the virus, the richness in nerves, and 
the extent of the injury to these nerves. 

The action of the virus of rabies on the nerve cen- 
ters and on the nerves is indirect ; this virus does not 
act as a chemical agent would, strychnine, for exam- 
ple. The anatomical changes, little visible to the naked 
eye, appear under the microscope as inflammatory 
lesions at different stages of their evolution. These 
lesions have their seat principally in the nerves of the 
bitten part and in the corresponding part of the spinal 
cord. They consist of congestions and capillary 
hemorrhages, with infiltration of leucocytes ; other 
lesions observed are limited foci of necrosis, diverse 
degenerations of the central nerve cells, transforma- 
tion of the myeline, hypertrophy of the axis cylin- 
ders, etc. 

The rabid symptoms are the expression of the neu- 
ritis and myelitis which successively develop. These 
changes manifest themselves by phenomena of excite- 
ment or of depression according as they are more or 
less advanced. Such phenomena are usually suc- 
cessive in occurrence, but may appear separately ; 
hence the two forms, furious rabies and dumb or 
paralytic rabies. Most frequently they are combined, 
and mixed forms result, intermediate between the two 
preceding, thus establishing the unity of the disease. 
The symptomatic tableau will naturally vary accord- 
ing to the part of the cerebro-spinal axis first invaded, 
and therefore according to the place of inoculation. 

The experiments of Pasteur, in which rabies was 



Microbic Diseases Individually Considered. 307 

transmitted by intra-venous inoculation, demonstrate 
the possibility of the absorption of the virus and its 
transmission by the circulatory fluids. 

Attenuation and Vaccination. The vjrus of rabies is 
not absolutely fixed; its passage through the organ- 
ism of the rabbit exalts its virulence, as we have seen 
above, whilst its inoculation to the ape enfeebles it. It 
becomes acclimated in the ape after three generations 
and then requires an incubation of twenty-three 
days, the incubation for natural rabies- being only 
eleven days. The fixed virus of the ape only rarely 
communicates the disease to the dog by intra-cranial 
inoculation, and intra-venous inoculation is always 
inoffensive ; this virus can be used to vaccinate the 
dog. 

Babes has shown that the virus of rabies introduced 
into the lymph sac of the frog becomes progressively 
attenuated until, after a certain time, it is capable of 
acting as a vaccine for the dog. The lymph of the 
frog removed from the body and mixed with an 
emulsion of a virulent cord also attenuates the ac- 
tivity of the latter. 

Attenuation by desiccation. A virulent rabic cord 
progressively loses its activity by desiccation in the 
air. This attenuation manifests itself by retarding 
the appearance of the disease, increasing the dura- 
tion of incubation ; thus, after two days' desiccation 
the fixed virus of the rabbit has not changed, and 
produces the disease in rabbits after seven days; after 
three to five days the incubation lasts for eight days; 
after six days it is extended to fourteen days ; after a 
desiccation of more than seven days the disease is no 
more communicated to the rabbit. 



308 Manual of Veterinary Microbiology. 

In order to obtain a regular and stable attenuation 
Pasteur starts with the cord of a rabbit which has 
been rendered rabid by means of fixed virus ; he di- 
vides it into segments of two centimeters in length 
and these he suspends in a wide-mouthed liter bot- 
tle containing pieces of caustic potash ; this bottle 
is placed in a chamber in which the temperature is 
maintained at 20 C. 

Variations in the temperature lead to modifications 
in the attenuation ; at 28 the virulence is destroyed 
in five days ; at 35 in four hours. 

The Russian rabbit, which has a smaller cord, al- 
lows of a more rapid attenuation. 

The temperature and the oxygen of the air are 
instrumental in producing this loss of pathogenic 
activity; the attenuation is slower in carbonic acid; 
the loss of humidity seems to have less influence; 
Protopopoff has, indeed, shown that this medication 
occurs also in glycerin-bouillon. 

Authorities are far from being unanimous as to 
the theory of this attenuation. Pasteur thinks that 
it consists merely in an impoverishment in active 
virus and not in a diminution of activity. 

The hypothesis of a chemical vaccine has been dis- 
pelled by Babes who showed that the rabic sub- 
stance filtered through porcelain, as well as that 
heated to 100, has no vaccinal property. Gamaleia 
has demonstrated that vaccinal injections have no. 
action on an individual already affected with the dis- 
ease, which also dispels the theory of a chemical vac- 
cine. Virus attenuated by desiccation regains the 
activity of the fixed virus on its second passage 
through the rabbit, 



Microbic Diseases Individually Considered. 309 

Anti-rabic vaccinations. It is possible to confer 
immunity against rabies both upon man and sus- 
ceptible animals. 

This immunity has been noted in certain cases of 
natural rabies in which recovery took place. It was 
especially well observed by Hoegyes in cases of ex- 
perimental rabies followed by recovery. This author 
records thirteen cases of recovery, six being of the 
furious form, out of one hundred and fifty-nine in- 
oculations to the dog. The immunity which followed 
these recoveries was still in force after five years. 

Most generally, immunity is observed after inocu- 
lations which are not followed by rabic symptoms ; 
it may be obtained in various ways. Pasteur ob- 
tained it in a dog by subcutaneous inoculation of 
street virus in large amount; this immunity, how- 
ever, is not the rule, and in some cases rabies itself 
appears. 

Hoegyes obtained immunity by injections of fixed 
virus diluted in 0*7 per cent aqueous salt solution. 
He made six inoculations in the dog with virus di- 
luted to 1 to 5,000, 1 to 2,000, 1 to 500, 1 to 100, and 
1 to 10. The animal showed no sickness and con- 
tracted a solid immunity against bites and diverse ex- 
perimental inoculations. 

Immunity was obtained by Pasteur by inoculating 
the dog with virus more or less attenuated in the 
ape. 

The same author has conferred immunity on the 
dog by subcutaneous inoculation of virus attenuated 
by the method of desiccation above described. The 
first inoculation is made with the cord of a rabbit 
desiccated during fourteen days, that is, one which 



310 Manual of Veterinary Microbiology, 

had lost virulence for the rabbit seven days before ; 
on the following days inoculations were made with 
cords which had been subjected to a less lengthened 
desiccation and, finally, with the virulent cord. The 
latter was inserted under the skin, in the blood, or in 
the cranial cavity ; some of these vaccinated rabbits 
were subjected to bites of rabid dogs; these differ- 
ent virulent inoculations produced no effects, and the 
persistence of the refractory state was verified several 
years later. 

Pasteurian vaccination. The period of incubation of 
of rabies after bites being quite lengthy, Pasteur con- 
ceived the idea of vaccinating, during this period, the 
subject which had received the bite, with the view of 
preventing the invasion of the disease. Experience 
has justified this illustrious savant in his belief. He 
has succeeded, by means of a series of dried cords, 
in vaccinating bitten dogs and even dogs which had 
been inoculated by trephining. It is evident that 
such preventive inoculations should be made as soon 
as possible in order that the refractory state be estab- 
lished before the active virus attains the nerve cen- 
ters. The results obtained in animals have served as 
a basis for the method of preventing rabies after bites 
in the human species, a method which is now applied 
in several establishments, and the beneficial results 
of which are no longer contested. Statistics show 
that the mortality from rabies in man has fallen from 
16 per cent to 0*67 per cent. 

Prevention of rabies after bites in ruminants. M. 
Galtier has shown that the intra-venous injection of 
ruminants with the virus of street rabies does not 
communicate the disease but confers immunity. This 



Microbic Diseases Individually Considered. 311 

author first, and later MM. Nocard and Roux, showed 
the efficacy of this method in the protection of 
ruminants recently bitten by rabid dogs. An emul- 
sion is made with virulent cord in sterilized water, 
strained through a linen cloth and injected into the 
jugular or ear vein. The injection, which should be 
of large amount, is not attended with danger. 

Typhoid fever of the horse.* 

Under the title of the typhoid affections several 
diseases were formally included which, clinically, had 
not been sufficiently differentiated. Our knowledge 
of these diseases is now somewhat more extended 
through the labors of Schiitz, Perroncito, Chante- 
messe and Delamotte, Cadeac, Galtier and Violet, 
etc. Many points are yet to be elucidated, but two 
diseases at least have been described which should 
not be included in the group of typhoid affections. 
These are pneumonia or contagious pleuro-pneu- 
monia, and pneumo-enteritis of fodders which will 
be considered later. 

Equine typhoid fever is especially characterized by 
stupor and great depression of the nervous and mus- 
cular systems of the affected animals; they are weak 
and unsteady on their feet, hold the head low, and 
move with a staggering gait as if under the influence 
of a profound intoxication or narcosis ; the eyes are 
weeping, half-closed, and often inflamed; the con- 
junctival mucous membrane is infiltrated, blood-shot, 

* [The term * typhoid fever," or " typhoid affections " of the 
horse appears to include the same diseases or complications of 
one disease which in England and America are described as In- 
fluenza, Epizootic cellulitis, etc. D.] 



Si 2 Manual of Veterinary Microbiology. 

and of a yellow tinge which may become yellowish- 
red when conjunctivitis supervenes. 

The fever of invasion is followed by diverse local- 
izations ; generally the digestive symptoms predomi- 
nate; the tongue is dry and more or less thickly 
coated, and symptoms of gastro-enteritis quickly 
appear. When the lungs become affected this com- 
plication occurs always several days after the begin- 
ning of the disease. The changes in this organ also 
differ absolutely from those which characterize in- 
fectious pneumonia. Typhoid fever predisposes to 
passive congestions; hence the lung becomes cedem- 
atous rather than hepatized and tubular breathing 
over the affected region is never heard, as in the case 
of pneumonia. This tendency to venous hypersemia 
shows itself in the limbs by cedernatous engorge- 
ments. 

All attempts to transmit the typhoid disease have 
been ineffective; the horse, ass, dog, and rabbit failed 
to contract th.e disease by the different methods of 
inoculation in common use. Equine typhoid, how- 
ever, conducts itself like a contagious disease; unlike 
contagious pneumonia it is polymorphic, showing 
itself sometimes as an enteritis, sometimes as a cardi- 
tis, and sometimes as a disease of the lungs, etc., 
whilst in a stable in which pneumonia prevails, all 
the affected horses show the latter lesion from the 
first. 

The formerly mooted question as to the identity of 
equine and human typhoid fever is now settled ; 
Eberth's bacillus does not occur in the equine disease 
and its inoculation to the horse remains without 
effect. 



Microbic Diseases Individually Considered. 313 

Contagious pneumonias of the horse.* 
The results obtained by different investigators, re- 
garding contagious equine pneumonia are far from 
being concordant. Nevertheless, it has been well 
established that we must separate from the typhoid 
affections one or more diseases which especially in- 
volve the lung. Whilst in typhoid fever the pulmo- 
nary localization is delayed, in infectious pneumonia 
the fever of invasion is of short duration and the 
disease localizes itself in the lung from the first. 
The early signs of depression in pneumonia are not 
to be compared with the state of prostration of ani- 
mals affected with typhoid fever, and these symptoms 
subside when hepatization is accomplished ; then the 
animals recover their accustomed liveliness. The 
conjunctival mucous membrane has a saffron red tint, 
the eye is bright and well opened. A rusty discharge 
from the nose appears during the first stage. 

The pulmonary inflammation may be lobar, occa- 
sionally lobular, or it may be complicated with pleu- 
risy (pleuro-pneumonia) ; other complications occa- 
sionally supervene, affecting the kidney, synovial 
membranes, articulations, the heart and its serous 
coverings, the meuinges, nerve centers, gastro-intes- 
tinal apparatus, etc. 

Galtier and Violet assert that the intestine is often 
affected along with the lung, even as a primary lesion 
(pneumo-enteritis), and that the disease also affects, 

* [In English veterinary works referred to as a complication of 
influenza. Ger. Brustseuche. D.] 

27 



314 Manual of Veterinary Microbiology. 

secondarily, the liver, spleen (it becomes enlarged 
and uneven on the surface), the bladder, articula- 
tions, tendinous sheaths, muscles, and keratogenous 
membrane. 

Schiitz has described, as the cause of this disease, 
an ovoid microbe, often associated in pairs and which 
possesses a capsule comparable with that of the 
pneumococcus of man. Perroncito has also observed 
a capsule, but states that it does not react to staining 
agents like that of the pneumococcus; further, the 
microbe studied by Perroncito kills the rabbit and 
the guinea pig, whilst the pneumococcus is inactive 
for the latter. Chantemesse and Delamotte attribute 
the disease to a streptococcus. Galtier and Violet 
describe two different microbes which gave two simi- 
lar diseases : a streptococcus and a diplococcus (strep- 
tococcus et diplococcus pneumo- enteritis equi). Cadeac 
found in all cases only one micrococcus, often grouped 
in pairs, sometimes in chains. 

In some investigations of our own, (1) we have found 

(1) In several horses affected with pneumonia we have found the 
streptococcus already described by Chantemesse and Delamotte 
and studied by Galtier. The elements of the chains are stained 
by the method of Gram and Weigert; cultures in bouillon pro- 
duce a flocculent precipitate ; gelatin is not fluidified. Inoculation 
in the rabbit causes a rapid emaciation and a severe diarrhoea, 
which results in death ; the latter is often preceded by pulmonary 
hemorrhage. At the autopsy the blood is dark, little plastic, and 
the serum deeply tinged by the coloring matter of the dissolved 
corpuscles ; the cavities of the peritoneum, pleura and pericar- 
dium always contain an abnormal quantity of reddish colored se- 
rum. The intestines are highly inflamed ; Peyer's patches are 
injected, sometimes studded with petechise ; the liver is discol- 
ored and tumefied; the spleen is enlarged, uneven, dark and fri- 
able; the kidneys show congestion and extravasation ; the pleura 



Microbic Diseases Individually Considered. 315 

the streptococcus of Chantemesse and Delamotte. 
The microbe, of Schiitz and those of Galtier and Vio- 
let do not take the Gram, whilst all the others do. 
Most of them are facultative anaerobes. All are 
pathogenic for the rabbit, which contracts a rapidly 
fatal disease : the blood is decomposed and the color- 
ing matter of the corpuscles, thus set free, passes 
into the serum, giving to the latter a red tinge, which is 
communicated to the liquids of the serous membranes 
and to the viscera in contact with these liquids ; the 

sometimes contains a fibrinous exudate ; the lung is much con- 
gested, this condition being sometimes complicated with intersti- 
tial hemorrhages ; the blood often escapes into the bronchi un- 
changed ; once only was hepatization present and that in a rabbit 
which had survived five days. 

The guinea pig is a very unreliable reagent for the streptococ- 
cus, whilst the dog is unaffected by it. 

Inoculation of 5 cc. of a culture into the lung of a glandered 
horse caused a very intense febrile reaction ; the temperature rose 
from 38'5 to 40*1 and remained at this figure during several 
days, whilst the animal showed complete inappetence and great 
prostration ; it usually kept the recumbent position and had to be 
assisted to its feet. It was impossible to detect any symptoms of 
pulmonary disease. At the autopsy, made ten days later, nothing 
was found at the place of inoculation, but four pneumonic centers 
existed on the lower border of the lobes of the organ. Three of 
these had the dimensions of a five franc piece, the fourth that of 
a child's hand. At these places the lung was very consistent, of a 
deep brown color, and manifestly hepatized. Two other horses 
received an injection of a five days old culture in the lung. At 
the autopsy, made on the following day in one case, and on the 
third day in the other, the place of inoculation showed an inflam- 
matory focus large as a man's fist ; the hepatized lung was dotted 
with very fine hemorrhagic points and the visceral pleura infil- 
trated with plastic serosity and notably thickened. In these le- 
sions the streptococcus of the cultures was found. None of these 
last cases showed any febrile reaction. 



316 Manual of Veterinary Microbiology. 

spleen is much enlarged. The diplococcus pneumo- 
enteritis equi has a less pronounced dissolving action 
on the blood corpuscles. Inoculation to the horse 
causes pneumonia; this disease develops as a conse- 
quence of injection into the lungs, trachea, or circu- 
lation, but only rarely as a result of ingestion. 

According to Gal tier and Violet, the disease origi- 
nates when horses are fed on forage of bad quality, 
such as soiled, moldy, or rusty hay, and imperfectly 
harvested and damaged grain. These damaged foods 
are the bearers of the germs of the disease and it is 
by their intermediation that these germs obtain entry 
to the system. The dust rising from the fodder en- 
ters the respiratory passages, where the microbes 
which it carries along act directly upon the lung. 
The disease has, in fact, been produced in the horse 
by inoculation of the products obtained by macera- 
tion of suspected foods. 

The disease can then be transmitted from one ani- 
mal to another by the excrements and nasal dis- 
charge; contagion, according to Galtier, plays an ac- 
cessory part; the diffusion of the cause suffices to 
explain the enzootic and even epizootic character of 
the affection. Its transmission, however, can not be 
doubted. 

Contagious pleuro-pneumonia of cattle. 
Microbe. M. Arloing found that the serosity 
which flows from the surface of a section through a 
diseased lung is very poor in microbes and that the 
majority of the culture bulbs inoculated with a small 
quantity of this serosity remain sterile. To obtain 
fertile cultures a large quantity of this serosity must 



Microbic Diseases Individually Considered. 317 

be transferred to the culture medium, or the inocu- 
lation made with the material obtained by scraping 
the surface of the section. By sowing the latter 
product on gelatin M. Arloing isolated four different 
species, one being a bacillus (pneumo-bacillus lique- 
faciens bovis) which rapidly fluidifies gelatin, and the 
other three micrococci. Of the latter, one produces 
white colonies resembling drops from a wax candle, 
the second gives whitish colonies which become 
wrinkled on aging, and the third, colonies which take 
an orange yellow tint. M. Arloing attributes the 
disease to the pneumo-bacillus liquefaciens bovis. The 
four microbes inoculated separately under the skin 
of a steer give an inflammatory tumefaction which 
disappears in five or six days ; the largest tumefaction 
is caused by the bacillus, and if several successive 
generations are inoculated it ultimately happens that 
the bacillus alone produces a local reaction. More- 
over, the bacillus alone is constantly present in dis- 
eased lungs. M. Arloing noticed that the isolated 
effects of the microbes which he had cultivated re- 
sembled only in a remote way those produced by 
fresh serosity, but he observed, also, that this last 
becomes more active in passing through the cellular 
tissue of healthy cattle. By taking the microbes 
from this reinforced serosity he obtained more viru- 
lent cultures of the pneumo-bacillus; 4cc. of cul- 
ture injected into the lung of one steer and 20 cc. 
injected into the veins of another produced the speci- 
fic lesions of pleuro-pneumonia. This author isolated 
from cultures a soluble substances which possesses 
remarkable phlogogenic properties, and which, of 
itself alone, reproduces the characteristic inflammatory 



318 Manual of Veterinary Microbiology. 

engorgements of the hypodermic inoculation of active 
virus. He attributes to this substance the inflamma- 
tions which sometimes occur, in the course of the 
natural disease, in points remote from the thorax. 

Whatever opinion may be held regarding these in- 
vestigations;, the virulent agent of pleuro-pneumonia 
exists in the serosity which flows in abundance when 
a diseased lung is incised. Inoculation of this liquid 
produces the following effects : 

In the subcutaneous cellular tissue a more or less in- 
tense inflammatory engorgement results ; in regions in 
which the connective tissue is loose and abundant 
this reaction often assumes a severe, progressive and 
occasionally gangrenous character, and leads to 
death ; the general effects of the virus are indicated 
by a febrile reaction of greater or less intensity. 
Pleuro-pneumonia is almost never observed as a con- 
sequence of subcutaneous insertions. But the latter 
confers immunity against later inoculations. 

The injection of the virus into the veins gives no 
more characteristic pulmonary lesions ; it confers 
immunity without any local manifestation, unless 
some of the virus should fall on the perivascular 
cellular tissue, in which case a dangerous tumor re- 
sults. M. Thiernesse, however, observed pleuro-pneu- 
monia after an injection of 35 grams in the jugular. 

Vaccination. In 1852 Willems recommended pre- 
ventive inoculation against this disease; his method 
was put in practice in different places and the pre- 
ventive action of Willemsian inoculation placed be- 
yond doubt. 

The inoculation may be performed in three princi- 
pal ways: by dermic, hypodermic, and intravenous 



Microbic Diseases Individually Considered. 319 

insertion. Derrnic insertion gives uncertain results, 
the virulent matter requiring to be brought into the 
cellular tissue in order to act efficaciously. On the 
other hand, hypodermic injection supplies the best 
conditions for the evolution of the virus. For the 
reception of the pulmonary serosity the cellular tis- 
sue of the tail should be selected in preference to all 
other regions; as has already been said the conse- 
quences of inoculation are often very severe in regions 
where the connective tissue is loose and abundant. 
In the tissue of the tail, one or two drops of the 
serosity are sufficient. It is well to make a second 
vaccination a few weeks later ; this may be made in 
an interdicted region although this procedure can not 
be recommended, especially when the reaction to the 
first inoculation has been insufficient. 

With the view of obviating the accidents which 
occasionally follow the caudal inoculation some ex- 
perimenters, including Professor-Director Degive, 
have had recourse to intravenous injection ; this 
method procures a solid immunity, but requires special 
care in order to avoid accidental contact of the virus 
with the cellular tissue which surrounds the vein, an 
accident which may result in a serious, perhaps fatal, 
engorgement. Larger doses may be introduced into 
the veins than into the cellular tissue. 

For the practice of Willemsian inoculation it is 
essential that fresh virus be employed. This is ob- 
tained by making a clean incision of a diseased lung 
with an aseptic knife and collecting the. serosity 
which flows spontaneously from regions in which the 
inflammation is most recent. As such a lung is not al- 



320 Manual of Veterinary Microbiology. 

ways at hand efforts have been made to preserve the 
virus. 

M. Laquerriere showed that in a frozen lung the 
virus remains intact for one year, at least. 

M. Nocard has quite recently recommended the 
preservation of the virus by the addition of half a 
volume of a five per cent solution of carbolic acid 
and half a volume of pure neutral glycerin. This 
mixture retains its virulence for months. 

For the purpose of procuring pure virus some' have 
advised its cultivation by direct inoculation in the 
cellular tissue of the calf and collecting the serosity 
of the inflammatory engorgement. 

Others again have sought for a means of mitigating 
the effects of the natural virus in order to diminish 
the accidents which result from its use; Pasteur has 
shown that it is preserved for six weeks in sealed 
tubes, but, at the same time, becomes so attenuated 
that it can be inoculated in an interdicted region 
without exciting fatal lesions. It has also been pro- 
posed to dilute the virus in water. Dilutions of 1 to 
50, 1 to 100, and 1 to 500 have yet sufficient activity 
to produce considerable inflammatory reactions and, 
therefore, to confer immunity. 

Septic pleuro-pneumonia' of calves. 
This disease occurs in an enzootic form on certain 
farms, attacking and quickly killing calves while still 
quite you ng. The lesions by which it is characterized 
have some analogy with those of contagious pleuro- 
pneumonia of larger cattle, but the thickening of the 
connective tissue septa is less marked and the flow 
of serosity less abundant, whilst the individual pul- 



Microbic Diseases Individually Considered. 321 

rnonary lobules have not the uniform color seen in 
the latter disease. We have observed that the pul- 
monary lesions begin, as in pleuro-pneumonia, in the 
interlobular connective tissue and, in the peripheral 
lobules, progress toward the center. Poels has 
noted, in calves affected with this disease, the fre- 
quency of sero-fibrinous exudates and pleuritic ad- 
hesions, lesions which have also been recorded by 
M. Yanden Maeghdenbergh. He also mentions the 
occasional occurrence of inflammations of the peri- 
cardium, liver, kidneys, stomach and intestine. 

Microscopical examination shows, in the lung and 
in the muco-pus of the bronchi, the presence of ovoid 
microbes with rounded ends, measuring from 1/z to 
l'5^e in length by 05// in thickness, easily stained by 
the aniline colors, but not stained by the Gram or 
Weigert methods. When stained with a very dilute 
aqueous solution of gentian violet they fix the color 
at their extremities, whilst the center remains clear. 
They are motile, vegetate rapidly in bouillon and on 
solid media, and are pathogenic for various species 
of animals. 

The rabbit dies in twenty-four to forty-eight 
hours after subcutaneous inoculation or ingestion of 
cultures or virulent products. In the same animal 
intra-pulmonary inoculation of a drop of culture pro- 
duces a pneumonia. Death is a little later in the 
guinea pig than in the rabbit. 

Two calves, aged thirteen days and eight weeks, 
and a one-year-old heifer, were inoculated by Poels, 
the first in the right pleura, the second in the trachea, 
and the third in the lung. They died after 20, 54, 
and 66 hours, respectively, with lesions of septic 



322 Manual of Veterinary Microbiology. 

pleuro-pneumonia. A pig also took the disease by 
pulmonary injection ; the sheep and the dog are re- 
fractory (Poels). The microbe is described as a facul- 
tative parasite capable of living in the soil, which 
fact, according to Poels, explains the persistence of 
the disease on an infected farm. M. Galtier thinks 
that the latter ought to be attributed to the mothers 
(see page 325). However that may be, its transmis- 
sion from calf to calf must be admitted ; the virulent 
germs are spread around with the expectorations and 
perhaps with the excrements (Poels claims to have 
found the germs in all the organs). These microbes, 
and especially those of the nasal discharge, con- 
taminate the vessels in which milk is fed to the dis- 
eased, and the same vessels are then used for other 
animals.* 

The microbe of this pleuro-pneumonia of calves 
belongs to the great class of ovoid bacteria showing 

*[In the diarrhoea or dysentery of young calves known as 
"white scours" (Ger. Kalberruhr) Jensen (1892) discovered in 
the blood, spleen, liver, kidneys and lungs, as well as in the mu- 
cous membrane of the intestine, oval bacteria, isolated or asso^ 
ciated in pairs or short chains, staining at the extremities only, 
and easily cultivated in the various artificial media. Bouillon 
cultures fed to young calves in doses of 5 cc. produced the charac- 
teristic diarrhoea and death in from one to two days. From the 
contents of the intestine of healthy calves he isolated apparently 
the same germ but found it destitute of pathogenic properties. 
Jensen came to the conclusion that the microbe is a facultative 
parasite, a usually harmless inhabitant of the intestinal canal but 
one which under certain abnormal conditions of the intestine 
(perhaps attributable to the diet) acquires pathogenic properties 
which become increased by subsequent passage from calf to 
calf. D.] 



Microbic Diseases Individually Considered. 323 

a clear central space, which are met with in a series of 
diseases. Hueppe groups these affections under the 
name of hemorrhagic septicaemias and attributes them 
all to the same germ. These diseases are : Koch's rab- 
bit septicsemia, fowl cholera, duck cholera, parrot dis- 
ease, infectious pneumonia of the pig, pneumo-enteritis 
of the pig, pneumo-enteritis of sheep, epizootic of wild 
game (Wildseuche), epizootic of ferrets, disease (bar- 
bone) of buffaloes, etc. It would be superfluous to in- 
sist on the non-identity of the germs of these different 
affections,but their close relationship can not be denied. 
The morphological characters of the germs are almost 
identical; all take the stain only at their extremities, 
leaving a clear space in the middle ; they do not stain 
by the Gram method, do not fluidify gelatin, and are 
pathogenic for rabbits. It is probable that several of 
the diseases mentioned above are due to one and the 
same microbe (fowl cholera and rabbit septicaemia, 
for example) and that all these micro-organisms rep- 
resent varieties of one fundamental species.* 

* [Billings found in specimens of the blood and organs of cattle 
which had died from the so-called "corn-stalk" or "corn-fodder 
disease " of the Western States an organism presenting great mor- 
phological resemblance to the germ of "swine plague" (hog 
cholera) but showing slight differences in its growth on the vari- 
ous culture media. It belongs to the group of ovoid, bi-polar stain- 
ing organisms and, according to this author, is the cause of the 
disease above named. The germ was found to be pathogenic for 
mice, rabbits and guinea pigs ; subcutaneous inoculation of a steer 
gave rise to fever, pneumonia and pleurisy, and great emaciation, 
terminating in recovery. From the original communications on 
this subject we learn that the germ is identical with a bacterium 
described by Burril as the cause of a disease of corn (maize) 
(Ills. Univ. Exper. Sta. Bui. 6), and the cattle become infected by 
feeding on the leaves of this diseased corn. Horses are also said 



324 Manual of Veterinary Microbiology. 

Epizootic abortion. 

Epizootic abortion is a contagious disease most fre- 
quently seen in the cow but also noticed in the ewe, 
she-goat, and even in the mare. The abortion occurs 
at all periods of gestation after the third month ; in 
the same animal it occurs at a later stage in each suc- 
ceeding year and at length allows the foetus to be 
carried till term, if, indeed, the cow, after a first 
abortion, has not become sterile. The calf is most 
frequently still-born ; in some cases it is born alive but 
its health is precarious; shortly after birth it emits a 
peculiar lowing; after the third day it is attacked 
with diarrhoea and death quickly follows. 

According to Nocard the disease takes its origin in 
diverse germs met with in the uterus of the animals 
which abort, germs which are never found in healthy 

to die from the same disease (Neb. Agric. Exper. Station, Bulletins 7, 
8, 9, 10). 

Noeard studied an " infectious broncho-pneumonia " which was 
observed in a number of recently imported American cattle (from 
Virginia, Indiana and Illinois). The lesions in the lung some- 
what resembled those of contagious pleuro-pneumonia. The 
muco-pus of the bronchi, the hepatized lung tissue, and the 
serosity contained a short, ovoid, motile bacterium, apparently in 
pure culture. It measured barely I/* in length by 0'3/* to 0*4^ in 
.thickness, stained with aqueous solutions of fuchsin and methy- 
lene blue, leaving a clear unstained central space ; not stained by 
the methods of Gram or Weigert. The germ was found to be 
pathogenic for the mouse, rabbit, guinea pig and pigeon, which 
die from subcutaneous inoculation; inoculated in the lung of 
calves and sheep it occasions a fatally-ending exudative broncho- 
pneumonia. Nocard believes this disease to be identical with 
the " corn fodder disease " of Billings. (Recueil de Med. Vet. Aug., 
1891. D.] 



Microbic Diseases Individually Considered. 325 

animals. These germs, among which is often found 
a micrococcus, isolated or in chains, are also present 
in the amniotic fluid, in the digestive canal of the 
aborted calves, as well as in the substance of the 
medulla oblongata of those individuals which during 
life gave utterance to the peculiar lowing sound just 
referred to. Nocard thinks that these germs give rise 
to a disease of the foetus and its envelopes, the mother 
remaining healthy. He explains the repeated abortion 
by the persistence of these germs in the womb, and 
the sterility by the acid reaction which they produce 
in the uterine secretions. 

The invasion of a stable by this disease generally 
coincides with the introduction of a pregnant infected 
cow; abortion then occurs annually in a certain num- 
ber of animals in this stable. The lengthy period of 
incubation of the disease implies an early infection. 

Galtier thinks that the disease is due to a general 
infection of the mother, which communicates the dis- 
ease to the foetus. Calves which are-not prematurely 
expelled will still harbor the germs, and these germs 
produce the pneumo-enteritis which occasions such 
ravages in these animals. 

In short, epizootic abortion appears to depend on 
multiple causes which are yet to be discovered, but 
in all cases the method of treatment laid down by 
Nocard is to be recommended. (1) 

(1) The prophylactic measures recommended are as follows: 

1. Each week the floor of the stable should be scraped, thor- 
oughly cleansed, and sprinkled with a solution of sulphate of 
copper : 40 grams to the litre. 

2. Each week from the date of conception the vagina of the 



326 Manual of Veterinary Microbiology. 



Contagious mammitis of milch cows. 
MM. Nocard and Mollereau have seen and de- 
scribed a special form of mammitis occurring in 
milch cows and readily passing from one animal to 
another. It appears in the form of indurated lumps 
which commence at the base of the teat, gradually in- 
crease in size, and may sooner or later invade the 
whole organ. The milk is diminished in quantity, 
becomes acid in reaction, and soon coagulates often 
as soon as it is drawn from the udder ; it is often 
mixed with pus, and grumous ; sometimes it exhales 
an offensive odor. These characters it communicates 
to good milk with which it may be mixed. 

Microbe. This is a rounded or ovoid micrococcus; 
it measures 1-25/z in length by I// in thickness and 
forms long straight or sinuous chains. It frequently 
appears bi-lobed, in way of division. It is aero- 
pregnant cows should be thoroughly injected by means of a large 
syringe, with the following tepid solution : 

Distilled water, ... 20 liters. 

36 per cent alcohol j ^ m ^ 

Glycerin, 

Bichloride of mercury, . . -10 grams. 

3. Each week, at the time of grooming, the vulva, anus, and 
lower surface of the tail of all the pregnant cows should be care- 
fully washed with a sponge saturated with the same tepid solu- 
tion. 

4. If a cow should abort it will be necessary to remove the 
placenta by hand, to destroy the foetus and after-birth by fire or 
boiling water, and to irrigate the uterus by means of a long tube 
introduced to the bottom of the cavity, with eight or ten liters of 
the tepid solution indicated above but containing only half the 
proportion of sublimate. 



Microbic Diseases Individually Considered. 327 

anaerobic. These characteristic chains are found in 
the milk and in the wall of the excretory ducts. 

Action of physical and chemical agents. The growth 
of the microbe in cultures is checked by a trace of 
boric acid; MM. Nocard and Mollereau, taking ad- 
vantage of this peculiarity, injected on several occa- 
sions at intervals of eight days 100 grams of a tepid 
four per cent aqueous solution of boric acid into the 
teats of the affected udders, the injection naturally 
being made immediately after milking. The parasite 
is also destroyed by a three per cent solution of car- 
bolic acid ; the authors recommend this solution for 
washing the hands of those who undertake the milk- 
ing. By these measures they succeeded in arresting 
the extension of the disease. 

Cultures. Alkaline bouillon with the addition of 
sugar or glycerin forms a medium well suited for its 
growth ; at 35 it forms a mass of very long chains 
which sometimes become agglomerated in silky flakes 
which after several days are deposited ; the reaction 
becomes acid in twenty-four to forty-eight hours; if 
chalk be added to the bouillon so as to neutralize the 
acid as it is produced, the culture is more vigorous 
and retains its vitality longer. Crystals of lactate of 
lime are often found at the bottom of the bulb. The 
growth of this microbe, therefore, gives rise to the 
lactic fermentation ; it can not, however, be identi- 
fied with the lactic bacillus. Cultures almost in- 
variably die after a few weeks. The microbe grows 
also on the different solid media. 

Experimental inoculations. Inoculation of pure cult- 
ures into the teat reproduced the disease in the cow 
and the goat; from the first day the inoculated udders 



328 Manual of Veterinary Microbiology. 

supplied a milk very rich in streptococci; that from 
the cow quickly showed an acid reaction and became 
clotted; the udder finally became inflamed. Inocu- 
lation in the mamma of a nursing bitch remained 
without effect; the dog, cat, rabbit, and guinea pig 
were also unaffected by intra- venous and in tra peri- 
toneal inoculations. 

Etiology. The disease is communicated through 
the intermediation of those who have charge of the 
milking of the cows, their hands being soiled with 
the diseased milk. When the latter is mixed with 
good milk this also takes the same characters. Con- 
tagion to the cows must, therefore, be prevented by 
disinfection of the hands, and contagion to the nor- 
mal milk by keeping the milk from the diseased cows 
in separate vessels. Such milk is, moreover, unfit for 
human consumption. 

Besides this disease, remarkable on account of its 
extreme contagiousness, the udder of the cow is sub- 
ject to various microbic lesions. M. Lucet especially 
called attention to infectious forms of mammitis due 
to an external cause; in a series of cases of acute 
mammitis he found one or several germs; these were 
sometimes micrococci, sometimes bacilli, and some- 
times both together. Penetration of the germs most 
frequently occurs through solutions of continuity of 
the integument of the udder. The microbe being 
different in different cases, we can readily under- 
stand that the severity of the disease will be very 
variable. 

Among the number of infectious forms of mam- 
mitis with internal cause tubercular mammitis should 
be especially mentioned. 



Microbic Diseases Individually Considered. 329 

Gangrenous mammitis of milch ewes. 
This disease, also called mal de pis, araignee, quickly 
kills ewes which are attacked by it ; it is due, accord- 
ing to the researches of M. Nocard, to a very fine mi- 
crococcus, measuring 0'2/z in diameter, and associated 
in groups of four or more, never in chains. It is 
stained by the method of Gram, is aero-anaerobic, 
and communicates to bouillon and to milk an acid 
reaction, coagulating the latter in twenty-four hours. 
Cultures retain their virulence only when renewed 
every day. When a culture is inoculated into the 
teat of a ewe it produces a rapidly fatal mammitis. 
The goat is refractory. The rabbit contracts an ab- 
scess from which it quickly recovers. The dog, cat, 
and guinea pig show only a local oedema. 

Diseases of milk. 

Milk just withdrawn from the udder is free from 
germs, but after its extraction it may quickly become 
infected and undergo a series of deteriorating changes. 
The most important of these deteriorations will here 
be briefly described. 

Curdled milk. The curdling of milk results from 
the lactic fermentation of milk sugar, the acid pro- 
duced then bringing about the coagulation of the 
casein. 

The usual cause of this fermentation is the bacte- 
rium lactis. This is a short, non-motile rod, measur- 
ing in length I// to 3// by O'6/i in thickness, most fre- 
quently isolated but occasionally arranged in series, 
and capable of spore formation. 
28 



330 Manual of Veterinary Microbiology. 

Other germs give rise to the same deterioration of 
milk, among these being the cocci of suppuration, 
erysipelas, contagious mammitis of the cow, etc. 
The infection occasionally originates in the udder 
(mammitis), more frequently, however, after the milk 
is withdrawn. The infection is facilitated by lack of 
cleanliness in the stables, dairy utensils, etc. 

The diseased milk coagulates more or less quickly 
according to circumstances; the process is hastened 
by heat. The cream separates imperfectly from such 
milk and the agglomeration of the butter globules is 
difficult. 

Putrid milk. Milk, like all organic liquids, readily 
putrefies. Various putrefactive germs are concerned 
in this process, but those which are most constant 
are the bacterium termo and lineola. 

The former is represented by short motile rods, 
measuring 1-4/* by 07//. The second species consists 
of large cylindrical, motile rods, measuring 3// to 5// 
by l-5/i. 

Infection results from lack of cleanliness ; the in- 
fected milk quickly putrefies, with the production of 
putrid gas in its substance. At the same time the 
cream takes a yellow color and a bitter or rancid 
taste, and butter can no longer be obtained from it; 
it then gradually disappears. 

Viscous milk. In this form of deterioration, which 
is observed one or two days after milking, the milk 
is not readily coagulable and the cream separates im- 
perfectly ; butter is obtained with difficulty and has a 
disagreeable taste. 

The disease is due to rounded elements 1^ in di- 
ameter, isolated or associated in the form of chains, 



Microbic Diseases Individually Considered. 331 

and transforming the lactose into a mucilaginous 
substance which gives to the milk its peculiar con- 
sistence. 

Blue milk. It sometimes happens that milk, from 
twenty to thirty-six hours old, shows on its surface 
small light-blue patches which, later, take an indigo- 
blue shade. These patches increase in area as well 
as in depth although never exceeding the thickness 
of the cream. Such milk is highly subject to change; 
it quickly becomes acid, coagulates and then putre- 
fies. 

The causative agent of blue milk is the bacillus 
cyanogenus. This is a motile rod, 2/i to 4// long by 
0'5// thick, commonly isolated but sometimes united 
in a zoogloea. The spore which it produces is a 
little larger than the bacillus giving to the latter a 
club or spindle-shaped appearance. In certain artifi- 
cial media it assumes very diverse involution forms- 
balloon and ribbon shapes, etc. Grown on the gela- 
tin plate it forms, after two days, small whitish spots 
which soon extend over the whole surface giving it 
a bluish color. Stab cultures in gelatin show whitish 
colonies on the surface and steel blue in the depth. 

When inoculated to milk it increases the alkalinity 
of the latter and the layer of cream becomes slate 
colored, this tint turning to blue on the addition of 
acid. If the milk has not been sterilized the lactic 
acid fermentation which goes on at the same time 
supplies the acid by which the blue color is pro- 
duced. 

The germ multiplies in albuminous solutions con- 
taining lactate of ammonia, without producing the 
coloring principle. 



332 Manual of Veterinary Microbiology. 

This coloring matter is most abundantly produced 
at about 20, in less amount at 25, and not at all at 
37^. The pigment is a soluble substance which gives 
a red reaction with potash, violet with ammonia. 
Inoculation of this germ to animals produces no re- 
sult. 

The infection of milk takes place after its with- 
drawal from the udder and is contingent upon a pre- 
vious infection of the dairy or of the stable. 

Cheese made from infected milk may also show 
this blue color, but the latter changes more or less 
to green in consequence of the yellow color which 
cheese assumes hi aging. 

Red milk. Two species of germs are capable of 
imparting a red color to milk. One, the micrococcus 
prodigiosus, is elliptical, motile, and forms on potato 
an abundant slimy growth of blood-red color. The 
infected milk shows on the surface a pellicle of a 
more or less deep red color, the deeper layers remain- 
ing unaltered. Another, the bacterium lactis erythro- 
genes, is a very short non-motile rod. In milk it 
slowly precipitates the casein and the whole mass as- 
sumes a blood-red color. 

Red milk is of much rarer occurrence than blue 
milk. 

Yellow milk. This deterioration occurs especially 
on boiled milk ; golden yellow patches appear on the 
milk which, at the same time, coagulates and becomes 
alkaline. The change is produced by the growth of 
the bacillus synxanthus, a slender, very mobile rod. 

Bacterial hcemoglobinuria of cattle. 
This acute febrile disease prevails in an endemic 



Microbic Diseases Individually Considered. 333 

form in certain marshy regions of Roumania, where 
it causes the loss of large numbers of oxen. Cows 
are less susceptible while calves seem to be refractory. 
It is characterized by the presence of albumen and 
haemoglobin in the urine ; the latter is red colored 
but does not contain blood corpuscles. The autopsy 
reveals the presence of interstitial extravasations and 
ulcers of the fourth stomach and duodenum. The 
tissue surrounding the kidneys is infiltrated with 
blood and serosity, the kidneys are friable and dark 
red in color and their pelvic mucous membrane 
ecchymosed; the bladder is filled with red colored 
urine. The liyer is tumefied and discolored, the 
spleen enlarged and darkened and the pulp diffluent. 

Babes has discovered in this disease a rounded mi- 
crobe 0.5// in diameter, usually arranged in pairs, 
sometimes in tetrads; it is decolorized by the Gram 
stain. It is found in the blood adhering to the red 
blood corpuscles or situated in their interior, but more 
especially in the serosity of the hemorrhagic cedemas 
and in the vessels of the kidney. It also exists in 
the vessels of the intestinal ulcers. The invaded red 
blood corpuscles are more or less altered. 

The rabbit, by inoculation of the blood or cedema- 
tous liquid, as well as by ingestion of the products 
of the disease or its cultures, contracts a general dis- 
ease which often terminates fatally. In the ox, the 
introduction into the veins or connective tissue of a 
considerable quantity of blood or of juice expressed 
from the kidneys, reproduces the typical disease with 
hsemoglobinuria.* 

* [In many respects similar to this haeinoglobinuria of cattle and 
to a closely allied disease of sheep (" Carceag ") also investigated 



334 Manual of Veterinary Microbiology. 

Distemper of young dogs. 

The contagiousness of this disease is well estab- 
lished although our knowledge of the germs which 

by Babs is the disease of cattle known in America as " Texas 
fever." It occurs in an acute and in a mild form, the latter be- 
ing in a sense endemic in the southern United States, whilst the 
acute form prevails in northern cattle which have been imported 
to these regions or which have been exposed to the infection 
brought north by cattle from the infected districts. The chronic 
or mild type, as it occurs in southern cattle, generally passes un- 
perceived. Mild forms of the disease are also frequent among 
northern cattle, especially when they receive the infection during 
the cooler weather of autumn. The acute disease (which occurs in 
the northern States only during the hot weather of summer) is 
generally fatal; in non-fatal cases it is sometimes followed by a 
prolonged period of unthrift and debility. Calves are less sus- 
ceptible than adult animals and after one or more attacks acquire 
a certain degree of immunity; to this cause is attributed the com- 
parative immunity possessed by cattle native to the southern 
States. It appears, however, that this immunity is not perma- 
nent and may be lost when such cattle are kept for several sum- 
mers in non-infected regions. The most noticeable symptoms of 
the acute disease are those of an intense continuous fever with 
the passage, in most cases, of red or dark-red colored urine ; un- 
steady gait and .muscular tremors in the neck and limbs occur 
toward the last stage ; occasionally symptoms of delirium are ob- 
served. 

Pathological anatomy. The most important lesions observed at 
the autopsy are : Injection of the vessels and occasionally patches 
of extravasation in the subcutaneous connective tissue ; blood 
thin and watery (Smith), or frequently of normal appearance; 
lungs normal or discolored by congestive patches ; heart muscle 
congested, points of extravasation on the pericardium and endo- 
cardium ; spleen enlarged, its capsule streaked and mottled by 
the injected vessels, pulp dark red and diffluent ; liver generally 
much enlarged and darkened from blood congestion or light yel- 
low in color from extreme engorgement with bile ; gall blad- 
der full of thick, dark, grumous bile ; kidneys generally congested, 



Microbie Diseases Individually -Considered. 335 

determine it is as yet incomplete ; some authors have 
described a bacillus ; others, a bacillus and a coccus; 

often showing irregular discolored patches beneath the capsula 
which itself is non-adherent, points of extravasation in the pelvic 
mucous membrane ; in many cases extensive effusion of bloody 
serosity in the fat and connective tissue surrounding one or both 
kidneys ; the urine contained in the bladder generally dark red 
in color free from blood corpuscles but strongly albuminous, 
sometimes of normal appearance ; congestion of the mucous folds 
of the fourth stomach ; more or less hypersemia and extravasation 
in the walls of the small intestine have also been noticed. 

Etiology. Texas fever in its acute and best known form occurs 
under two conditions, 1st, when .northern cattle are shipped into 
southern infected regions, and second, when cattle from perma- 
nently infected regions, brought north during the summer months, 
infect pastures in which susceptible cattle are subsequently al- 
lowed to graze. In the first case the disease may, and frequently 
does, appear two weeks after exposure to infection, in the second 
a lapse of six weeks or more intervenes between the arrival of the 
infection-bearing cattle and the outbreak of the disease. These 
differences are attributable to the method by which the infec- 
tion is conveyed into the bodies of susceptible animals. The in- 
vestigations of Smith and Kilborne (afterward repeated with the 
same results by others) have shown .that this takes place through 
the intermediation of cattle ticks (BoophiluB bovis, Curtice), the 
progeny of those adherent to the skin of the southern cattle. 
The lapse of time between the arrival of these cattle and the out- 
break of the disease represents the time required for the incuba- 
tion of the next generation of these parasites along with the pe- 
riod of incubation of the disease. In the case of cattle shipped 
into permanently infected districts their invasion by ticks and 
consequent infection may begin at once. It is possible also that in 
permanently infected districts infection may occur by other means. 
Later generations of ticks which come to development on sus- 
ceptible cattle in the North are also capable of communicating 
the disease. Horses, which may also be invaded by these para- 
sites, do not obtain the disease, and the progeny of ticks which 
have developed on these animals have not been shown to be dan- 
gerous for cattle. 

Experimental inoculations. Positive results have been obtained 



336 



Manual of Veterinary Microbiology. 



and others, again, attribute the disease to a micro- 
coccus. We have found as the result of our own 

|}y Smith by intra-venous and subcutaneous injection of suscep- 
tible cattle with the blood of animals suffering from the disease 
as well as with the blood of apparently healthy southern cattle. 
No other animal species has been found to be susceptible. Direct 
contagion seems, however, rarely or never to occur under natural 
conditions. 

Microbe. Cover glass preparations from the blood, spleen, liver, 
kidney, and heart muscle of cattle which have died from acute 
Texas fever, show the presence of a rounded or somewhat ovoid, 
or pyriform (Smith) body, isolated, in pairs, or occasionally three 
or four, within a certain proportion of the red blo'od corpuscles. 
They are usually extremely abundant in the juice of the kidney 
(where they also occur between the cellular elements), less so in 
the liver and spleen, and still less in blood from the large ves- 
sels or heart. These stain readily with aqueous solutions of ani- 
line colors (methylene blue), as well as with haematoxylin (Dela- 
field's). Their outline after staining is generally less well defined 



Fig. 




Smear preparation from kidney of ox. 
Texas Fever. X1000. (D.) 

al, led to negative results. 



than bacteria 
similarly stain- 
ed ; neverthe- 
less their ap- 
pearance with- 
in the unstain- 
ed disk of the 
red blood cells is 
quite character- 
istic and in the 
absence of other 
evidence is of 
itself sufficient 
for a diagnosis 
of the disease. 

Culture tests of 
the blood and 
organs in cases 
of Texas fever 
have, in gener- 
Billings obtained an ovoid " bi-polar " 



Acute 



Microbic Diseases Individually Considered. 337 

researches, bearing upon four subjects only, that 
a micrococcus in pure culture is always present 
in the pustules of the cutaneous form. M. Mathis 
has recorded in a special work the results of his in- 
vestigations. He succeeded in demonstrating in the 
contents of pustules, in the discharge from the nos- 
trils, in the blood and in the tissues, the presence of 
spherical micrococci, isolated or grouped in pairs, 
in chains, or in masses ; these measure 0*l/z to O3// in 
diameter. He cultivated these organisms in bouil- 
lon; this medium becomes turbid, then, after fifteen 
to twenty days, clears again by the deposition of the 
suspended germs at the bottom of the vessel. 

Subcutaneous inoculation of these cultures in sus- 
ceptible dogs is followed by an cedematous tumefac- 
tion, with pustules on the skin in the region of the 
inoculation ; generally there is an elevation of tem- 
perature and occasionally generalization of the pus- 
tulous eruption, with cough, discharge from the nose, 
etc., and if young subjects are experimented with, 
the disease may terminate fatally. 

staining bacterium, pathogenic for small animals, which he de- 
scribes as the cause of the disease. Smith, to whose investiga- 
tions our knowledge of the etiology of Texas fever is chiefly due, 
regards the intra-corpuscular body already mentioned, which he 
discovered in 1889, as the causative agent of this disease. He de- 
scribes it as occurring in several forms representing different 
stages in its development ; the parasite, according to this author, 
belongs to the protozoa (Pyrosonaa bigeminum). The destruction 
of red blood corpuscles, the essential characteristic of this disease, 
is brought about by the direct action of the parasite. The latter 
exhibits amoeboid motion within the corpuscle which it ultimately 
destroys and then is found in its free stage between the cellular 
elements. Attempts at cultivation were unsuccessful. D.l 
29 



338 Manual of Veterinary Microbiology. 

The experimental disease confers immunity. M. 
Mathis has found that immunity also follows the 
natural disease. 

Notwithstanding the importance of the results al- 
ready obtained concerning the etiology of distemper 
of dogs, there are some obscure points which yet re- 
quire to be elucidated, such as, for instance, the inter- 
esting question of the pathogenesis of those nervous 
troubles which so frequently complicate the disease. 

Phosphorescent meats. 

Dead animal matters not unfrequently become 
phosphorescent. Marine fishes and mollusks are 
especially liable to be thus affected, while meats are 
less subject to this change. Phosphorescent meat 
shows on its surface a coating which is luminous in 
the dark, and easily removed by scraping. 

The cause of this deterioration resides in a bacte- 
rium, \p. in length (Photobacterium Pflugerii). This 
germ grows well on meat and fish, especially at tem- 
peratures between 10 and 30. In presence of oxy- 
gen it gives to the culture media a whitish glimmer. 
The invasion of meat takes place very quickly in 
summer, one piece of meat being readily infected by 
another. The multiplication of the germ is rather 
favored by salting, and it ceases when putrefaction 
begins. The luminous property is directly connected 
with the life of the germ. It has not hitherto been 
found that such meats are poisonous. 

In order to complete this subject we should yet 
have to discuss a number of diseases the microbic 
nature of which is unquestionable. Thus, cocci and 



Microbic Diseases Individually Considered. 339 

streptococci have been described in the vesicles of 
aphthous fever, bacilli in rinderpest, micrococci in 
cow-pox or vaccinia, as well as in enzootic hepatitis 
of young pigs, etc. But not only is there a lack of 
unanimity as to the morphology of the germs of 
these diseases, but the study of their special biologi- 
cal characters has yet entirely to be made, hence, a 
particular acquaintance with these germs is only of 
secondary importance to the practitioner. 



340 Appendix. 



APPENDIX. 

(FROM ATTFIELD'S CHEMISTRY.) 

The Metric System of weights and measures is 
founded on the meter. The engraving represents a 
pocket folding measure, the tenth part of a meter in 
length, divided into ten centimeters, and each centi- 
meter into ten millimeters : 

Fig. 19. 



The Decimeter. 

The units of the system with their multiples and 
submultiples are as follows : 

UNITS. 

Length. The Unit of Length is the METER, derived 
from the measurement of the quadrant of a meridian 
of the earth. (Practically, it is the length of certain 
carefully-preserved bars of metal from which copies 
have been taken.) 

Surface. The Unity of Surface is the ARE, which is 
the square of ten meters. 

Capacity. The Unity of Capacity is the LITER, 
which is the cube of a tenth part of a meter. 

Weight. The Unit of Weight is the GRAMME, which 
is the weight of that quantity of distilled water, at 






Appendix. 



341 



its maximum density (4 C.), which fills a cube of the 
one-hundredth part of the meter. 



TABLE. 

Note. Multiples are denoted by the Greek words " Deca," Ten, 
" Hecto," Hundred, " Kilo," Thousand. 

Subdivisions, by the Latin words, " Deci," One-tenth, "Centi," 
One-hundredth, " Milli," One-thousandth. 



QUANTITIES. 


LENGTH 


SURFACE. 


CAPACITY. 


WEIGHT. 


1000 
100 
10 
1 (Units) 
.1 
01 
.001 


Kilo-meter 
Hectometer 
Deca-meter 
METER 
Deci-meter 
Centi-meter 
Milli-meter 


Hectare 
ARE 
Centiare 


Kilo-liter 
Hecto-liter 
Deca-liter 
LITER 
Deci-liter 
Centi-liter 
Milli-liter 


Kilo-gramme 
Hecto-gramme 
Deca-Gramme 
GRAMME 
Deci-gramme 
Centi-gramme 
Milli-gramme 



Relation of Metric to United States measures of 
Length, Capacity, and Weight : 



1 Meter, . 
1 Decimeter, 
1 Centimeter, 
1 Millimeter, 



39. 370432 inches. 
3.937043 " 
0.393704 " 
0.039370 " 



Approximately, 1 centimeter (cm.) = in.; 1 millimeter (mm.) 
^5- in.; or conversely, 1 in. = 2 cm. = 25 mm. 
In micrometry, 1 Micron (/u) = 0.001 mm. == 0.00004 in. = 



(Milliliter) 1 cubic centimeter (ccm.) = 16.23 minims. 
(Liter) 1,000 cubic centimeters = 33.81 fluidounces. 

Approximately, 1 liter = 1 quart. 

1 Gramme (weight of 1 ccm. of water at 4 C.) = 15.432 grains. 
1 Kilogramme (1,000 grammes) = 15432.350 grains. 

Approximately, 1 kilogramme = 2 Ibs., Avoir. 



342 



Appendix. 



Therm-ometric scales. On the Centigrade (C.) scale 
the freezing point of water is made zero, and the 
boiling point 100 ; on the Fahrenheit (F.) scale the 
zero is placed 32 degrees below the congealing point 
of water, the boiling point of which becomes, conse- 
quently, 212. 

The degrees of one scale are easily converted into those 
of another if their relations be remembered namely : 
180 (F.), 100 (C.) : that is, 18 to 10, or 9 to 5. 

Formula? for the Conversion of Degrees of one Ther- 
mometric Scale into those of another : 

F = Fahrenheit ; C = Centigrade ; D = The observed degree. 

If above the freezing point of water (32 F.; 0.CO, 

FintoC, . . . (D 32) -- 9X5. 

CintoF, . . . D -T-5X9 + 32. 

If below freezing, but above F ( 1777 C.), 
FintoC, . . (32 D) -*- 9 X 5. 

CintoF, . . . 32 (D-J-5X9. 

If below F ( 17 '77 0.), 

FintoC, . . . (D+32)-*-9X5. 

CintoF, . . (D-s- 5X9) 32. 

Equivalents on the Centegrade and Fahrenheit 
scales : 



C 


F 


C 


F 


C 


F 


C 


F 


C 


F 


10 


14 


25 


77 


37 


98-60 


44 


111 20 


75 


167 





32 


28 


82 40 


38 


100 40 


45 


113 


80 


176 


+8 


41 


30 


86 


39 


102 20 


50 


122 


90 


194 


10 


50 


32 


89 60 


40 


104 


56 


131 


100 


212 


15 


59 


34 


93 20 


41 


105 80 


60 


140 


120 


248 


20 


68 


35 


95 


42 


107-60 


65 


149 


150 


302 


22 


71-60 


36 


96 80 


43 


109 40 


70 


158 


200 


392 







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